Cary Winuv: Software Manual

Cary WinUV
Software Manual
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2 Cary WinUV Software Manual

Contents
Contents
1. Introduction 7
Applications 7
User documentation 10
Conventions 10
2. Installation 11
Computer requirements 11
Preparation 13
New computer 14
Upgrade 17
Starting the software 19
3. Software Overview 21
ADL Shell 22
Advanced Reads 22
Align 22
Color 23
Concentration 24
Enzyme Kinetics 24
GLP Administration 24
Kinetics 25
RNA-DNA Estimation 25
Cary WinUV Software Manual 3

Contents
Scan 26
Scanning Kinetics 26
Simple Reads 26
Spectroscopy Configuration Manager 26
System Information 27
Thermal (not for Cary 50) 28
UV Dissolution (Cary 50 only) 28
UV Fiber Optic Dissolution (Cary 50 only) 28
Validate 29
Varian Spectroscopy Database Administrator 29
File name extensions 29
Interface 31
Hint text 32
Using the Help 32
Navigating 33
Searching 34
Tracking „favorite‟ topics 35
Printing 35
4. How To… 37
Start an application 38
Manually read samples using Advanced Reads 39
Perform a calibration and manually measure concentrations using
Concentration 42
Enzyme Kinetics 48

Contents
Perform a temperature-controlled Enzyme Kinetics run

(Cary 50) 48
Perform a temperature-controlled Enzyme Kinetics run using
the Multicell Holder (Cary 100–6000i) 52
Kinetics 59
Perform a temperature-controlled Kinetics run (Cary 50)
59
Perform a temperature-controlled Kinetics run using the
Multicell Holder (Cary 100–6000i) 63
Perform a multi-wavelength, single cell, single rate Kinetics
run (Cary 100–6000i) 68
Perform an RNA-DNA run using the Multicell Holder (Cary
50) 73
Perform a temperature-controlled RNA-DNA run using the
Multicell Holder (Cary 100–6000i) 77
Scan 82
Perform a scan with baseline correction (Cary 50) 82
Perform a scan with baseline correction (Cary 100–6000i)
86
Perform a scan in Signal-To-Noise mode (Cary 100-6000i)
90
Perform a scan in Independent mode (Cary 5000) 95
Collect data (Cary 50) 99
Collect data using the Multicell Holder with temperature
control (Cary 100–6000i) 104
Perform a Simple Reads measurement at a single wavelength 110
Thermal 111

Contents
Perform a run and determine Tm using derivative

calculations 111
Perform a run and determine Tm using hyperchromicity
calculations 116
Perform a UV Dissolution run (Cary 50) 124
Perform a UV Fiber Optic Dissolution run (Cary 50) 138
Set up tests for validation 150
5. Troubleshooting 153
Instrument offline 154
Connect button instead of Start 154
Not enough memory 155
Poor display of videos and photographs 155
GLP log does not list some privilege changes 156
Report header and footer not being updated 156

Introduction
1. Introduction
Applications 7
User documentation 10
Applications
Agilent Cary 50/100/300/4000/5000/6000i instruments run using the
easy-to-use Agilent Cary WinUV software. The Cary WinUV software
consists of various applications, depending on the Cary package
ordered (refer to Table 1). A brief description of each application is
provided in Table 2. For detailed information about each application,
refer to Chapter 3.
NOTE Throughout this manual, UV Dissolution and UV Fiber Optic Dissolution users
should replace „Cary WinUV Software‟ with „UV Dissolution Software‟ or „UV
Fiber Optic Dissolution Software‟ respectively.

Introduction
Table 1. Applications included with each Cary WinUV software package

Package Applications
Analysis ADL Shell, Advanced Reads, Align, Concentration, GLP Administration, Kinetics,
Scan, Scanning Kinetics, Simple Reads, System Information, Validate
Bio ADL Shell, Advanced Reads, Align, Concentration, Enzyme Kinetics, GLP
Administration, Kinetics, RNA-DNA Estimation, Scan, Scanning Kinetics, Simple
Reads, System Information, Thermal (not for Cary 50), Validate
Color Color
Pharma ADL Shell, Advanced Reads, Align, Concentration, Enzyme Kinetics, GLP
Administration (not for CFR systems), Kinetics, RNA-DNA Estimation, Scan,
Scanning Kinetics, Simple Reads, System Information, Thermal (not for Cary
50), Validate, Spectroscopy Configuration Manager (administrator use only),
Varian Spectroscopy Database Administrator (administrator use only)
UV Dissolution ADL Shell, Advanced Reads, Align, Concentration, Enzyme Kinetics, GLP
Administration (not for CFR systems), Kinetics, Scan, Scanning Kinetics, Simple
Reads, System Information, UV Dissolution, Validate, Spectroscopy
Configuration Manager (administrator use only), Varian Spectroscopy Database
Administrator (administrator use only)
UV Fiber Optic Dissolution ADL Shell, Advanced Reads, Align, Concentration, Enzyme Kinetics, Kinetics,
GLP Administration (not for CFR systems), Scan, Scanning Kinetics, Simple
Reads, System Information, UV Fiber Optic Dissolution, Validate, Spectroscopy
Configuration Manager (administrator use only), Varian Spectroscopy Database
Administrator (administrator use only)

Introduction
Table 2. Descriptions of each Cary WinUV software application

Application Description Page
ADL Shell A pre-defined template for writing ADL programs. 22
Advanced Reads For collecting absorbance readings for multiple samples at single and 22
multi wavelengths (up to 6).
Align For aligning various instrument lamps and accessories. 22
Color For calculating color coordinates, determining color difference and 23
graphically displaying both.
Concentration For quantitative analysis. 24
Enzyme Kinetics For determining various parameters of enzyme activity at single and multi 24
wavelengths (up to 6).
GLP Administration For restricting operator access, password-protecting each application. 24
Kinetics For performing kinetics determinations at single and multi wavelengths 25
(up to 6).
RNA-DNA Estimation For collecting absorbance readings of nucleic acids. 25
Scan For running and view data collections. 26
Scanning Kinetics For performing kinetics determinations for a wavelength range. 26
Simple Reads For taking absorbance readings at a single and multi wavelength (up to 26
6).
Spectroscopy For administrators to set 21 CFR Part 11 security and access privileges in 26
Configuration Manager Pharma software.
System Information For recording your company and instrument details for use in reports, 27
and so on.
Thermal For temperature-based multicell measurements, typically DNA melts at 28
single and multi wavelengths (up to 6) (not for Cary 50).
UV Dissolution For performing tablet dissolution measurements (Cary 50 only). 28
UV Fiber Optic For monitoring tablet dissolution using fiber optics (Cary 50 only). 28
Dissolution
Validate For testing your instrument to ensure that it is working correctly. 29
Varian Spectroscopy Provides a database environment for storing and maintaining your data. 29
Database Administrator

Introduction
User documentation
You have been provided with the following documentation to help set
up and operate your Cary system:
 Installation Guides (Cary 50 only), with information on
unpacking the instrument, installing the interface card in the
computer and setting up the system.
 A hardware manual, with safety practices and hazards
information, instructions for installing and maintaining the
components of the Cary and troubleshooting information.
 This software manual, with instructions for installing the Cary
WinUV software, an overview of the software detailed ‘How To…’
procedures and software-related troubleshooting information.
 Extensive Help (provided with the Cary WinUV software)
containing context-sensitive Help, step-by-step instructions for
frequently performed analyses and instructions for using any
accessories you ordered. Refer to ‘Using the Help’ on Page 19.
Conventions
The following conventions are used throughout procedures in the
documentation:
 Menus, menu items, buttons and check boxes have been typed in
bold. For example, ‘click OK’ and ‘From the Edit menu, choose
Copy’.
 ALL CAPITALS indicate keyboard commands. For example,
‘Press ENTER’ and ‘Press SHIFT+F3’.
A Note is used to give advice or information.
A Tip is used to give practical hints to help you achieve the best
possible performance from your instrument.

Installation
2. Installation
Computer requirements 11
Preparation 13
New computer 14
Upgrade 17
Starting the software 19
This chapter assumes you have already set up the Cary instrument
and computer, including installing the instrument interface card in
the computer (if you supplied your own computer) and connecting
the instrument and/or computer to the power supply as described in
the appropriate Cary hardware documentation.
Computer requirements
Follow this recommended computer configuration for running Cary
WinUV software when buying a new computer:
 IBM compatible
 Intel® Pentium® IV processor (or later)
 At least 1GB RAM
 At least 10 GB free space on hard drive
 Video card supporting 800 x 600 pixel resolution, high color (16
bit) mode (or better)
 Super VGA screen (or better)
 20 x CD drive
 16 bit sound card
 Windows® 101 key keyboard

Installation
 Microsoft® or compatible mouse

 RS232 serial port
 One comm port if using the Agilent SPS 3 Sample Preparation
System in conjunction with the Internal Routine Sampler
 Two comm ports if using the Agilent SPS 3 Sample Preparation
System in conjunction with the External Sipper accessory
 Windows XP with Service Pack 3
 Microsoft Internet Explorer version 6* or later
 PCI slot for IEEE or Cary 50 PCI
* The Cary WinUV software uses functionality provided by Microsoft
Internet Explorer version 6.0. You do not need to use this as your
Web browser. If your company rules prevent the installation of
Internet Explorer version 6, you can use another browser, with some
loss in functionality.
Cary 50 only:
 One spare computer power supply connector
 Power supply rating of 220 W
 One spare slot for the accessory cable connector
 Room for a second connector from the PCI interface card

Installation
Preparation
Before installing the Cary WinUV software, ensure that you have:
Preparation requirement Reference

Installed Microsoft Windows XP operating system with Windows operating system documentation.
Service Pack 3 and checked that all devices, for example,
sound card and CD drive, are working.
Installed Windows Internet Explorer version 6 or later. Documentation supplied with Internet Explorer.
Set the monitor screen resolution to at least 800 x 600 Windows operating system documentation.
pixels and the color quality to at least High Quality.
Logged on as an Administrator. Windows operating system documentation.
Read the Late Breaking News. Any Late Breaking News documents delivered
with the Cary WinUV software.
NOTE The Cary WinUV software will read Cary OS/2 and DOS data files. However, if
you are upgrading to version 4.10 from Cary OS/2 or DOS, contact your local
Agilent office, as a service call will be required.
NOTE If you are installing 21 CFR Part 11 software, refer to the:

 Cary WinUV Pharma Software Installation Instructions for 21 CFR Part
11 Environments, publication number 8510258100
 UV Dissolution/UV Fiber Optics Dissolution Software Installation
Instructions for 21 CFR Part 11 Environments, publication number
8510237400
21 CFR Part 11 software installation involves an administrator using the Agilent
Spectroscopy Configuration Manager (SCM) to set up access privileges, and so
on. In addition, SCM uses the Varian Spectroscopy Database Administration
(VSDA) program for storing files. Refer to the SCM and VSDA Help for
information about setting up 21 CFR Part 11 systems.

Installation
New computer
To install the Cary WinUV software on a new computer:
1 Log on to the instrument computer with Administrator
privileges.
2 Insert the application software disk, select the preferred
language and click OK.
3 Follow the prompts on the screen until the ‘Select Destination
Location’ window appears. Confirm the directory in which you
would like to install the application. Alternatively, click Browse
to choose a different location. Click Next.
NOTE Agilent strongly recommends that the Cary WinUV folder and applications be
installed in the recommended C:\Program Files directory.
4 The ‘Folder Does Not Exist’ dialog box may appear. Click Yes to
create the folder.
5 Follow the prompts on the screen until the ‘Ready to Install’
window appears. Click Install.
NOTE During the installation of the .Net Framework and GPIB driver, the computer may
appear frozen and the „Cancel‟ button is unavailable. This is correct. The
installation can take 3 to 5 minutes. Do not try to exit the installation during this
time.
6 If prompted to complete installation of Cary WinUV by restarting

your computer, select No, I will restart the computer later. Click
Finish.
7 ‘Software Registration’ will be displayed. Click Next.

Installation
NOTE Ensure the software registration is completed by the User of the Cary UV-Vis-NIR
spectrophotometer. For further information, refer to the Agilent Software
Registration Instructions, publication number 8510256100.
8 Complete all the fields on the ‘Customer Details’ page. Click

Next.
NOTE The Product Key is found on the front of the Cary WinUV Software disk case that
was delivered with the instrument.
9 Complete all the fields on the ‘Product Details’ page. Click Next.
10 Complete all the fields on the ‘Work Environment Details’ page.
Click Register.
11 A dialog box appears stating ‘Your Agilent registration has been
successful’. If your computer is not connected to the Internet,
refer to the Agilent Software Registration Instructions,
publication number 8510256100 for more information.
12 ‘Validate Setup Wizard’ will be displayed. Click Next to install
the Validate version 4.10 (build 467) patch.
13 Remove the application disk from the CD drive.
14 Insert the disk labeled Help.
15 A list of Help files to be installed will be displayed. Click OK.
16 Follow the instructions on the screen to install the Help.
17 When the status indicates ‘Finished’, click Close.
18 Remove the Help disk from the CD drive.

Installation
NOTE At the end of the application software installation, you will need to turn off the
computer in order to install the computer-instrument interface card and to
complete the automatic detection and installation of the driver for the instrument
interface card. Refer to the Cary 50 Installation Instructions (publication number
8510185100) or the Cary 100/300/4000/5000/6000i Spectrophotometers
Hardware Manual (publication number 8510197200).
19 Once the card has been installed, turn on the computer. If using a
Cary 50, proceed to Step 20. If using a Cary
100/300/4000/5000/6000i, proceed to Step 25.
20 The instrument interface card should automatically be detected.
A ‘Found New Hardware Wizard’ dialog box will be displayed.
21 Select No, not this time and then click Next.
22 When prompted, select Install software automatically
(Recommended).
23 During installation of the hardware drivers for the Cary 50 PCI
card, a warning message will be displayed. Click Continue
Anyway.
24 Click Finish.
25 Select the System Information application from the Cary WinUV
folder on the Windows desktop. Enter the company details,
instrument type and serial number. Click OK.
26 Restart the computer to complete the installation of the
instrument interface card.

Installation
Upgrade
NOTE If you are upgrading 21 CFR Part 11 software, refer to the:
 Cary WinUV Pharma Software Installation Instructions for 21 CFR Part

11 Environments, publication number 8510258100.
 UV Dissolution/UV Fiber Optics Dissolution Software Installation
Instructions for 21 CFR Part 11 Environments, publication number
8510237400.
To upgrade the Cary WinUV software:

1 Cary 50 only: Turn off the computer and disconnect the Cary 50
interface cable from the instrument.
2 Log on to the instrument computer with Administrator
privileges.
3 If upgrading Cary WinUV version 3.1 or earlier, proceed to Step
4. If upgrading Cary WinUV version 4.10 (build 464), proceed to
Step 6.
4 Click Start > Settings > Control Panel > Add/Remove
Programs.
5 Click NI-488.2.1.60 and then click Change/Remove and follow
the prompts. When prompted click Yes to remove all unused
components.
6 From the Windows toolbar, right-click on the System
Information icon and click Shut Down System Information.
7 Insert the application software disk, select the preferred
language and click OK.
8 Follow the prompts on the screen until the ‘Select Destination
Location’ window appears. Confirm the directory in which you
originally installed the application software. If required, click
Browse to navigate to the correct location. Click Next.
9 The ‘Folder Exists’ dialog box will be displayed. Click Yes.

Installation
10 Follow the prompts on the screen until the ‘Ready to Install’

window appears. Click Install.
NOTE During installation of the Microsoft .Net Framework and GPIB driver, the
computer may appear frozen and the „Cancel‟ button is unavailable. This is
correct. The installation can take 3 to 5 minutes. Do not try to exit the installation
during this time.
11 If prompted to complete installation of Cary WinUV by restarting

your computer, select No, I will restart the computer later. Click
Finish.
12 ‘Software Registration’ will be displayed. Click Next.
NOTE Ensure the software registration is completed by the User of the Cary UV-Vis-NIR
spectrophotometer. For further information, refer to the Agilent Software
Registration Instructions, publication number 8510256100.
13 Complete all the fields on the ‘Customer Details’ page. Click

Next.
NOTE The Product Key is found on the front of the Cary WinUV Software disk case that
was delivered with the instrument.
14 Complete all the fields on the ‘Product Details’ page. Click Next.
15 Complete all the fields on the ‘Work Environment Details’ page.
Click Register.
16 A dialog box appears stating ‘Your Agilent registration has been
successful’. If your computer is not connected to the Internet,
refer to the Agilent Software Registration Instructions,
publication number 8510256100 for more information.
17 ‘Validate Setup Wizard’ will be displayed. Click Next to install
the Validate version 4.10 (build 467) patch.
18 Remove the application disk from the CD drive.

Installation
19 Insert the disk labeled Help.

20 A list of Help files to be installed will be displayed. Click OK.
21 Follow the instructions displayed on the screen to install the
Help.
22 When the status indicates ‘Finished’, click Close.
23 Remove the Help disk from the CD drive.
24 Select the System Information application from the Cary WinUV
folder on the Windows desktop; enter the Company details,
Instrument type and Instrument Serial number. Click OK.
25 If using a Cary 100/300/4000/5000/6000i, restart the computer to
complete the installation.
If using a Cary 50, turn off the computer and then connect the
Cary 50 interface cable to the instrument to complete the
upgrade.
Starting the software

To start the Cary WinUV software:
1 Click the Windows Start button then (All) Programs, Agilent
and Cary WinUV. Alternatively, double-click the Cary WinUV
folder on the desktop.
UV Dissolution/UV Fiber Optic Dissolution:
Click the Windows Start button, then (All) Programs, Agilent,
UV Dissolution or UV FO Dissolution. Alternatively, double-
click the UV Dissolution or UV FO Dissolution folder on the
desktop.
2 Select the desired application. Refer to Chapter 3 for information
about the available applications.
After the initial Cary flash screen appears, the application will open.
TIP To familiarize yourself with the Cary WinUV software, browse the Help after
installing the software. See „Using the Help‟ on Page 19.

Installation
This page is intentionally left blank.

Software Overview
3. Software Overview
ADL Shell 22
Advanced Reads 22
Align 22
Color 23
Concentration 24
Enzyme Kinetics 24
GLP Administration 24
Kinetics 25
Scan 26
Simple Reads 26
Spectroscopy Configuration Manager 26
System Information 27
Thermal (not for Cary 50) 28
UV Dissolution (Cary 50 only) 28
UV Fiber Optic Dissolution (Cary 50 only) 28
Validate 29
Varian Spectroscopy Database Administrator 29
File name extensions 29
Interface 31
Using the Help 32

Software Overview
This chapter provides a brief introduction to the Cary WinUV

software and the individual applications to help you familiarize
yourself with its use. For more detailed information about the
applications and their settings, refer to the extensive Help (see Page
19 for information about using the Help).
ADL Shell
The Applications Development Language (ADL) is a built-in
spectroscopy language supplied with the Cary WinUV software. The
ADL Shell gives you a pre-defined template for writing ADL
programs. Rather than needing to write the code for basic functions
such as graphing and filing, the ADL Shell has a number of these
commands already implemented. In other words, you do not have to
design your own interface — you can use the ADL Shell to provide the
basic functionality and build on that.
Advanced Reads
The Advanced Reads application can be used to read multiple
samples in a single run at a single or multi wavelength (up to 6). You
can use various accessories to take multiple sample solution and
aliquot readings in absorbance, percent transmittance, Abs*F or
percent reflectance mode, and find the mean.
Align
The Align application is used to align the lamp(s) in the instrument
and various accessories. It enables you to set instrument parameters
such as the beam mode and wavelength. It also allows you to keep
track of operating hours by lamp type.
The ‘Lamps’ page of the Alignment window allows you to monitor the
energy of the lamp(s). Instrument parameters can be changed on the
‘Cary’ page.
Align can also be used to configure the Series 1 and Series II 6 x 6
and 8 x 6 cell changer (Cary 100/300), the cell changer for microcells
(Cary 50/100/300/4000/5000/6000i), and the Series I and II Rear
Beam Attenuator (Cary 100/300/4000/5000/6000i).

Software Overview
Color
The Color application is used to measure and calculate the
transmitted or reflected color of samples. The following extensive
color calculations are supported:
 Tristimulus
 Chromaticity
 CIE Lab
 CIE LUV
 Hunter Lab
 Whiteness
 Yellowness (ASTM E 313-00)
 Tint (ASTM E 313-00)
 Gardner Color (ASTM D 6166-97 and DIN EN1557)
 Haze (ASTM D1003-61)
 Choice of 21 illuminants, including 2 user-specified illuminants
 Choice of 6 observers, including 2 user-specified observers
 Calculation interval of 1, 5 or 10 nm
 Thickness correction to calculate the color of the same sample in
varying thicknesses, rather than having to measure the samples
 Graphical display of color spaces
 Color difference calculations — Delta E, FMC-2, CMC and BFD
 Color matching tolerance circle using DE Lab

Software Overview
Concentration
The Concentration application is used to calibrate the system for
quantitative analysis. You can select from several fit types for your
calibration: linear, direct linear and quadratic. Based on the fit type,
the Concentration application will calculate the coefficients of the fit
equation and the correlation coefficient. Alternatively, you can define
your own equation for the calibration.
Enzyme Kinetics
The Enzyme Kinetics application uses Michaelis-Menten principles to
calculate the maximum rate (Vmax) and substrate concentration that
gives half the maximum rate (Km) of enzyme-catalyzed reactions.
Accurately obtaining Vmax and Km, requires you to perform numerous
Enzyme Kinetics runs at different substrate or inhibitor
concentrations to create a series of absorbance versus time curves.
The software determines the initial velocity (V0) of each curve, then
you enter the substrate and inhibitor concentrations for each cuvette.
The software uses the V0, [S] and [I] values to plot traces
representing absorbance versus time curves with a common [S] or
[I]. It is from these graphs that the software determines Vmax and Km.
Enzyme kinetics measurements can be performed at single or multi
GLP Administration
The GLP Administration application is used to protect the system
from unauthorized use, enabling application-specific privileges to be
turned on or off by the system administrator.
If this application is operational, users will need to be registered,
have a user name and a valid password before they can access the
various privileges.

Software Overview
NOTE GLP Administration is not present in UV Dissolution or UV Fiber Optic Dissolution

software. The Spectroscopy Configuration Manager included in that package is
used to provide good laboratory practice functions.
Kinetics
The Kinetics application is used to obtain absorbance versus time
data to enable you to determine the rate of reaction. Kinetics
measurements can be performed at single or multi wavelengths (up
to 6).
The Kinetics application allows:
 Calculation of Zero Order, First Order and Second Order reaction
rates from absorbance versus time data.
 Entry of activity factors for multiple cells.
 Overlay of the best-fit line on raw data.
 Auto or manual estimates for the first order and second order
Marquardt fitting.
RNA-DNA Estimation
The RNA-DNA Estimation application is used to calculate the
following parameters, used in determining the amount, type and
purity of nucleic acid samples:
 Absorbance of samples at selected wavelengths.
 A(260)/A(280) ratios with or without background correction at
320 nm.
 Absorbance ratios at your own nominated wavelengths with or
without background correction.
 Average ratio values for replicate samples.
 Protein and nucleic acid concentrations using Warburg Christian
coefficients.
 260 nm Factor Parameters, A(260) * F.

Software Overview
Scan
The Scan application enables you to set up and run wavelength
scans, with the collected scans displayed in the ‘Scan’ window.
Scanning Kinetics
The Scanning Kinetics application allows you to perform cyclic scans
across a wavelength or wavenumber range. From the resultant
absorbance versus wavelength data, an absorbance versus time
(kinetics) curve can be obtained for any wavelength in the range. The
kinetics curves can then be used to calculate Zero Order, First Order
and Second Order reaction rates.
You can choose an automatic or manual estimate for the First Order
and Second Order Marquardt fitting.
This application also enables you to correct samples for a baseline
during the scan. You can choose a 100%T baseline, or you can select
from other baseline options such as a zero baseline correction that
will apply both a 100%T and a 0%T baseline correction to your sample
scans.
Simple Reads
The Simple Reads application is used to perform quick absorbance
readings of samples at single wavelengths. To take a reading, click
‘Zero’ to zero the instrument and then click ‘Read’.
Simple Reads measurements can be performed at single or multi
Spectroscopy Configuration Manager

The Spectroscopy Configuration Manager (SCM) provides the system
administrator with a tool to manage your 21 CFR Part 11
environment. SCM provides the means to create, configure and
maintain data in relation to system security, user management and
data paths.

Software Overview
The Privileges and Profiles software controls which

applications/functions may be run by a particular user. It also
establishes the level of authority a user may have with regard to
signatures and accessing certain parts of an application
Agilent uses the SCM for security and permission rights. These
security functions provide:
 Access controls and authority checks via the use of user
identification codes and passwords.
 Electronic record security via the use of databases.
 Time and date stamped audit trails.
The use of user identification codes and passwords enables control
over who can log on to the system and who can perform particular
functions within the Agilent application software. It also provides the
mechanism to allow electronic signing of electronic records. The use
of databases coupled with SCM, prevents all unauthorized users from
changing or deleting files. The SCM event logs augment the audit
trails resident in the application software. The SCM administrator
must set up the required users. It is important that a number of
simple requirements are followed when this is done to ensure that
compliance with the 21 CFR Part 11 rule is maintained.
System Information
The System Information application allows you to enter company
and instrument information, and specify headers, footers and bitmap
or icon files to be used in reports (for example, your company logo).
System Information also allows you to specify the default settings for
the ‘Hint’ text, such as the Hint Pause (the length of the delay before
the hint appears when the pointer is moved over a control) and Hint
Hide Pause (the time the hint stays visible for if the pointer is not
moved off the control). These settings will be used by the other Cary
WinUV applications.

Software Overview
Thermal (not for Cary 50)

The Thermal application is used to perform thermal DNA analysis,
enabling you to calculate Tm using the various thermoelectric Cary
accessories. Once the data is collected, you can choose to calculate
the Tm value by either the derivative or hyperchromicity methods.
Thermal measurements can be performed at single or multi
UV Dissolution (Cary 50 only)

The UV Dissolution application is used to monitor tablet dissolution,
using up to two dissolution baths. The application enables you to
collect data for up to 10,000 minutes (8 days) at defined time
intervals.
The data can be displayed as absorbance, percent dissolved versus
time or milligrams dissolved versus time. You can then calculate the
time taken to reach a series of nominated percent dissolved values
and/or calculate the percent dissolution of the tablets at given times.
UV Fiber Optic Dissolution (Cary 50 only)

The UV Fiber Optic Dissolution application is used to monitor tablet
dissolution, using up to two dissolution baths and fiber optics. The
application enables you to collect data for up to 10,000 minutes
(8 days) at defined time intervals.
The data can be displayed as absorbance, percent dissolved versus
time or milligrams dissolved versus time. You can then calculate the
time taken to reach a series of nominated percent dissolved values
and/or calculate the percent dissolution of the tablets at given times.

Software Overview
Validate
The Validate application enables you to carry out a number of
performance tests to verify that the system is performing according
to specification. Included are the validation tests needed to satisfy
the requirements of the British Pharmacopoeia, European
Pharmacopoeia, US Pharmacopoeia and Therapeutic Goods
Association of Australia.
An optional Validation package is also available from Agilent,
providing detailed information on the functional specification and
development process of the Cary system. It also contains detailed
DQ/IQ/OQ documentation to assist your initial and on-going
validation activities.
Varian Spectroscopy Database Administrator

The Varian Spectroscopy Database Administrator (VSDA) is designed
for system administrators to set up and maintain the databases that
are used by the application software to store data. VSDA uses
Microsoft SQL Server 2005 for database operations.
Companies can use VSDA together with the Agilent Spectroscopy
Configuration Manager (SCM), the application software and their
own Standard Operating Practices to form a 21 CFR Part 11 capable
environment for controlling their Agilent instruments.
VSDA allows the data collected by Agilent instruments to be stored
locally (on the same computer as the application software), or
remotely in a Client/Server arrangement.
Configuration must be performed by the system administrator, or a
person with administration rights to run VSDA.
File name extensions

The various Cary WinUV software files, such as method files, report
files and graphic files are saved with a three-letter file name
extension that represents the type of file, and the application that
created the file.

Software Overview
The file name extensions for each file type are:
File type File name extension

Methods .m**
Data .d**
Report .r**
Graph Template .g**
Settings .s**
Baseline .c**
Batch .b**
ASCII .csv
Rich Text Format .rtf
Cary OS/2 .dat
Cary DOS data.
Grams .spc
ADL .adl
where ** equals the Cary WinUV application code, used to distinguish

between different applications. The application codes are:
Application File name extension

Advanced Reads .*ab
Color .*cl
Concentration .*cn
Enzyme Kinetics .*ek
Fabric Protection .*fp
Kinetics .*kn
RNA-DNA .*dn
Scan .*sw
Scanning Kinetics .*sk
Simple Reads .*sr
Sunglasses .*sg
Thermal .*tm
UV Dissolution/UV Fiber Optic Dissolution .*dt
Validate .*vo

Software Overview
For example, a method file from the Concentration application would

have the file name extension ‘*.mcn’.
Interface
A typical Cary WinUV software screen consists of a Menu line (‘A’ in
Figure 1), Instrument buttons (B), Command buttons (C), Toolbar
(D), Graph area (E), Report area (F) and Status line (G).
Figure 1. A typical Cary WinUV software screen

A. Menu line B. Instrument buttons C. Command buttons D. Toolbar
E. Graph area F. Report area G. Status line

Software Overview
Hint text
You can obtain ‘Hint text’ for a particular control or field by
positioning the pointer over the control/field name for a short time.
After a short delay, a brief hint will be displayed describing what the
control or field does. If you hold the pointer over a numeric entry
field, the valid range for that field will be displayed.
If these hints do not appear, choose ‘Hints’ from the ‘View’ menu to
enable them. Conversely, if the hint text is enabled and you wish to
turn it off, choose ‘Hints’ from the ‘View’ menu.
NOTE You can alter the properties of the hints, such as the length of the delay before
hints appear, on the „Hints‟ page in the System Information application.
Using the Help

The Cary WinUV software includes extensive Help, which should be
your primary source of information on how to use your Cary system.
It contains descriptions of the various application windows, dialog
boxes and fields that make up the software, as well as step-by-step
instructions to help you perform various tasks. It also includes ‘Tips
and Tricks', ‘Troubleshooting' and ‘Contacts' sections in case you
encounter difficulties using the software.
Since the Help system is so extensive, it is advisable to familiarize
yourself with the contents of the Help by viewing the Help Home
page. There are a number of ways to open the Help Home page:
 Click the Windows ‘Start’ button, ‘(All) Programs’, ‘Agilent’, ‘Cary
WinUV’, ‘Cary Help’.
Click the Windows ‘Start’ button, then ‘(All) Programs’, ‘Agilent,
UV Dissolution’ or ‘UV FO Dissolution’.
 Double-click the ‘Cary WinUV’ folder on the desktop and double-
click ‘Cary Help’.

Software Overview

Double-click the ‘UV Dissolution’ or ‘UV FO Dissolution’ folder on
the desktop and double-click ‘Cary Help’.
 Click ‘Help Topics’ on the ‘Help’ menu when in an application to
go to the Help Home page.
You can also make the Help open on the appropriate page containing
information relevant to the currently open application or dialog box:
 Click ‘(Application) Help’ (for example, ‘Scan Help’) on the ‘Help’
menu when in an application to go to the Home page for that
application.
 Click the ‘Help’ button on a dialog box to go to information about
that dialog box.
 Press F1 to go to information about the current dialog box.
Navigating
You can move around the Help using the ‘Contents’ on the left of the
Help window.
NOTE To display the Contents, click the „Show‟ button at the top of the Help window.
The Contents sections are accessed by clicking the icons and

associated text:
 To expand/contract a section in the Contents list, double-click
the ‘folder’ icon for that section, or click the ‘+’/‘-’ symbol next to
the icon.
 To display a topic, click the text associated with that topic.
The topics in the Help are often hyperlinked to other related
information. To return to the previous topic after clicking a
hyperlink, click the ‘Back’ button at the top of the Help window.

Software Overview
Searching
You can quickly search the Help system for specific information
using key words.
To search for information on a particular subject:
1 Click the Search tab (next to the ‘Contents’ tab).
2 Type the word(s) you want to search for and click the List Topics
button or press ENTER to list all the relevant pages containing
the search word/s.
The number of topics found will be displayed. You can view the
topics in alphabetical order by clicking ‘Title’ at the top of the list, or
in the order they appear in the Help by clicking ‘Rank’.
3 Select the desired topic from the list by highlighting it then
clicking the Display button. Alternatively, double-click the topic.
All occurrences of the keyword will be highlighted.
4 If this does not provide the information you require, enter a more
specific word or additional words in the key words field and try
again. Click the right arrow symbol to the right of the keywords
field and select any of these words AND, OR, NEAR and NOT to
place between your keywords.
TIP Another way to narrow your search is to use the check boxes at the bottom of
the Search page. For example, selecting „Search titles only‟ will only list those
Help topics containing the key word in the title.
Within a page
You can skip to a section of interest in a Help page by searching for a
relevant word.
To find a key word within a Help page:
1 Click anywhere on the Help page.
2 Press CONTROL+F to open the ‘Find’ dialog box.
3 In the ‘Find what:’ field, enter the word you wish to look for.

Software Overview
4 If required, limit the Find by selecting an option such as ‘Match

case’. (This option will only find the words that appear in the
same letter case (for example, all lower case) as the entered key
word.
5 Select the Direction of the Find. Select ‘Up’ to search for the key
word from the bottom of the page upwards, or ‘Down’ to start the
Find from the top of a page.
6 Click the Find Next button or press ENTER to begin the Find.
The first occurrence of the designated word will be highlighted.
7 Click the Find Next button or press ENTER repeatedly to jump to
other occurrences of the key word. When all instances of the
word have been found, the message ‘Finished searching the
document’ will be displayed.
Tracking „favorite‟ topics

You can keep a list of useful Help topics using the ‘Favorites’ option.
If you come across a topic you may wish to refer to again:
1 Click the Favorites tab. The title of the page currently visible will
be present in the ‘Current topic:’ field at the bottom of this tab.
2 Click the Add button to include this topic to the Favorites list.
If you would like to view this page later on, you can simply click the
‘Favorites’ tab and highlight the topic of interest by clicking it, then
click the ‘Display’ button. (Alternatively, double-click the topic.)
If a topic is no longer required in the Favorites list, highlight it then
click the ‘Remove’ button.
Printing
You can print one or more Help topics for a particular application.
To obtain a hard copy of the current Help page:
1 Click the Print button at the top of the Help window. The ‘Print’
dialog box will be displayed.
2 Click Print.

Software Overview
To obtain a hard copy of multiple Help pages:

1 Right-click on the topic in the ‘Contents’ page, and choose Print.
2 Click Print the selected heading and all sub-topics.

How To…
4. How To…
Start an application 38
Manually read samples using Advanced Reads 39
Perform a calibration and manually measure
concentrations using Concentration 42
Enzyme Kinetics 48
Kinetics 59
Scan 82
Perform a Simple Reads measurement at a single
wavelength 110
Thermal 111
Perform a UV Dissolution run (Cary 50) 124
Perform a UV Fiber Optic Dissolution run (Cary 50) 138
Set up tests for validation 150
This chapter provides step-by-step instructions on how to perform

common operating procedures using your Cary instrument and
various software applications. For more detailed information on the
software applications or your Cary instrument and accessories, refer
to the extensive Help (see Page 19 for information on using the Help).

How To…
Start an application
1 For all Cary WinUV applications except UV Dissolution/UV
Fiber Optic Dissolution:
Cary WinUV and click the application that you wish to run.
Alternatively, double-click the application icon in the ‘Cary
WinUV’ folder on the desktop.
For UV Dissolution/UV Fiber Optic Dissolution:
UV Dissolution or UV FO Dissolution and click the application
that you wish to run.
2 Select your instrument type if necessary and click OK to open the
application.
NOTE If you are running a GLP system, you will be prompted to enter a password
before accessing the application.
NOTE If you are using 21 CFR Part 11 software, you will be prompted to enter a user
identification, select the appropriate Group and Project and enter a password
before accessing the application.

How To…
Manually read samples using Advanced Reads

This procedure describes how to read samples in the Advanced
Reads application using no accessories.
1 Set up data collection parameters
Setup dialog box
In Advanced Reads, click the Setup button or choose Setup from the
Menu line to display the ‘Setup’ dialog box and specify the method
parameters for a new method.
2 Set up instrument parameters
Setup dialog box | Cary page
a In the ‘Wavelength’ field, enter the relevant wavelength.
b In the ‘Ave. (averaging) Time’ field, enter the required value.
A good starting point is 0.1 seconds.
c In the ‘SBW’ field, enter the required spectral bandwidth.
Unless your method specified another value, use the
maximum setting. (Not for Cary 50.)
d Select ‘Replicates’ or ‘Sample Averaging’. For Replicates,
enter the number of replicates of each sample that you would
like read. For Sample Averaging, enter ‘2’ for duplicate
aliquots of the sample, and so on.
NOTE If using a microcell, select a smaller spectral bandwidth.
e Select the ordinate mode you require from the drop-down list
in the ‘Y Mode’ field. Enter a ‘Factor’ value if you have
selected ‘Abs*F’.
3 Set up lamp options
Setup dialog box | Options page
If you are using a Cary 50, proceed to Step 4.

How To…
a Select Auto lamps off to automatically turn off the lamps at

the end of the collect. This option is especially useful when
performing reads overnight or unattended for long periods of
time. In some Cary instrument models, you may also choose
to use a third lamp, such as a mercury lamp (if installed).
b Click the UV/Vis button to use both lamps.
c Enter the wavelength at which you would like the source
lamp to change from the ultraviolet to the visible/near-
infrared lamp. The recommended changeover is
350 nanometers for lamps with an ultraviolet cutoff.
If you are using a Cary 5000/6000i, also enter the detector
and grating changeover wavelengths (800 nanometers is
recommended for both).
d Under ‘Beam Mode’, select the beam mode that you require.
In most cases this should be Double Beam and Normal. If
you select ‘Single Beam’ you also need to enter a value in the
‘Energy’ field.
4 Ensure no accessories are selected
Setup dialog box | Accessories pages
Make sure no accessories are selected on the ‘Accessories 1’ and
‘Accessories 2’ pages.
5 Set up your samples
Setup dialog box | Samples page
a Enter the ‘Number of Samples’. The table below expands or
contracts to match your choice.
b In the ‘Samples’ table, enter the name of each sample. You
can enter up to 20 characters for each name.
If the samples have the same name with a different numeric
extension, enter the name in the first sample position and
then click the ‘Increment’ button.
You can click the ‘Import’ button to use the names of samples
stored in a text file.

How To…
6 Set up reporting and printing requirements

Setup dialog box | Reports page
a Enter your name in the ‘Name’ field.
b Enter a comment relating to your experiment in the
‘Comment’ field.
c Set up your report style by selecting the appropriate check
boxes under ‘Options’. For example, select ‘AutoPrint’ to
automatically obtain a printout of your report. Select
‘Parameters’ to include your experimental parameters in the
report.
NOTE If „AutoPrint‟ is selected, the system will send the report information to the
specified printer as well as the Report area. However if „AutoPrint‟ is not
selected, the report will only be sent to the Report area.
7 Set up storage of collected data

Setup dialog box | Auto Store page
Select Storage off. The method, collected data and report will not be
automatically saved. However, you can manually save it all at the end
of the collection.
8 Set up visual system monitoring
Select Show Status Display on any of the Setup pages, or from the
View menu, to display information about your current reaction.
9 Finish setup
Once you are satisfied with your method setup, click OK to confirm
any changes you have made and close the ‘Setup’ dialog box.
10 Zero the instrument
a Click Zero. Alternatively, choose Zero from the Commands
menu.
b Place a blank in the sample compartment and click OK.

How To…
11 Read samples
a Click the Start button, press F9 or choose Start from the
Commands menu. The ‘Sample Selection’ dialog box will be
displayed.
b Select the samples you would like to read, then click OK.
c The ‘Present Sample’ dialog box will prompt you to place the
appropriate sample in the sample compartment. Click OK to
read the sample.
d Repeat for the remaining samples.
12 Save your data
a On the File menu, click Save Data As.
b Enter the ‘File name’ for this Concentration run.
c Click Save. The data will be stored as a Batch file.
13 Export your data
a On the File menu, click Export report (*.csv).
b Enter the ‘File name’ for this read.
c Click Save. The data will be stored as an ASCII spreadsheet,
with a *.csv file name extension.
Perform a calibration and manually measure concentrations using

Concentration
This procedure describes how to perform a multi-standard
calibration and measure sample concentrations in the Concentration
application using no accessories.
Setup dialog box
In Concentration, click the Setup button or choose Setup from the
a In the ‘Wavelength’ field, enter the relevant wavelength.

How To…
b In the ‘Ave. (averaging) Time’ field, enter the required value.

A good starting point is 0.1 second.
c Enter the required spectral bandwidth in the ‘SBW’ field. Use
the maximum setting unless your method specifies another
value. (Not for Cary 50.)
NOTE If using a microcell, select a smaller spectral bandwidth.
d Select ‘Replicates’ or ‘Sample Averaging’. For Replicates,

enter the number of replicates of each sample that you would
like read. For Sample Averaging, enter ‘2’ for duplicate
aliquots of the sample, and so on.
e Select the ‘Y mode’ you require. Click ‘Abs’ to specify
Absorbance mode or ‘Emission’ if you are measuring
fluorescence. (Not for Cary 50.)
f Enter an upper range and lower range value in the ‘Y min.’
and ‘Y max.’ fields to specify the displayed ordinate range.
These are starting values only. The Cary WinUV software will
automatically rescale the calibration graph as the standards
are measured.
3 Set up the calibration
Setup dialog box | Standards page
a Click the Standards tab to set up the standards and their
parameters associated with the data collection.
b Select Calibrate During Run to perform a calibration when
the ‘Start’ button is clicked.
c Set the appropriate units for your standards for reporting
purposes.
d Set the ‘Standards’ field to the number of standards that you
are using. The table below will expand or contract to match
your choice.
e In the ‘Standards’ table, enter the concentration of each
standard in the ‘Conc.’ column.
f Under ‘Fit Type’, select the type of curve fitting required for
your calibration.

How To…
g Enter the required R2 value or correlation coefficient in the

‘Min R2’ field. The closer the number is to 1.000, the better
the fit. Typically, 0.95 is used.
4 Set up lamp options
If you are using a Cary 50, proceed to Step 5.
a Select Auto Lamps Off if you want to automatically turn off
the lamps at the end of the collect. This option is especially
useful when performing Concentration runs overnight or
unattended for long periods of time.
b Click the UV/Vis button to use both lamps.
c Enter the wavelength at which you would like the source
lamp to change from the ultraviolet to the visible/near-
infrared lamp. The recommended changeover is
350 nanometers for lamps with an ultraviolet cutoff. If you
are using a Cary 5000/6000i, also enter the detector and
grating changeover wavelengths (800 nanometers is
recommended for both).
d Under ‘Beam Mode’, select the beam mode that you require.
In most cases this should be Double Beam and Normal. If
you select ‘Single Beam’, you also need to enter a value in the
‘Energy’ field.
Setup dialog box | Accessories page
Click the Accessories tab and make sure that no accessories are
selected.
a Enter the number of samples in the ‘Number of Samples’
field. The table below expands or contracts to match your
choice.
b In the ‘Samples’ table, enter the name of each sample. You
can enter up to 20 characters for each name.

How To…

You can click the ‘Import’ button to use the names of samples
stored in a text file.
automatically obtain a printout of your report.
specified printer as well as the Report area. However, if „AutoPrint‟ is not
selected, the report will only be sent to the Report area and can be viewed by
choosing „Report‟ from the „View‟ menu.
8 Set up weight and volume correction

a Under ‘Weight/Volume Corrections’, select Corrections to
activate the correction facility.
b Enter the theoretical sample weight in the ‘Method Weight’
field. This is the weight of the sample specified in your
method.
c Enter the weight units in the ‘Units’ field.
d Enter the theoretical sample volume in the ‘Method Volume’
field. This is the volume to which the method tells you to
make the sample.
e Enter the volume units in the ‘Units’ field.
f In the ‘Samples’ table, enter the actual weight and volume for
each sample.

How To…

Select Storage off. The method, collected data and report will not be
automatically saved. However, you can manually save it all at the end
of the collection.
11 Finish setup
a Click Zero. Alternatively, from the Commands menu, choose
Zero.
b Place a blank in the sample compartment and click OK.
13 Perform the calibration
a Click the Start button or, from the Commands menu, choose
Start. The ‘Standard/Sample Selection’ dialog box will be
displayed.
b Select the standards and samples to be used in the analysis.
By default all standards and samples are selected.
c Click OK.
d The ‘Present Standard’ dialog box will prompt you to place
the appropriate standard in the sample compartment. Click
OK to measure the standard.
e Repeat until you have measured all of the standards. The
Cary will calculate the calibration and the correlation
coefficient.

How To…
NOTE If the set correlation coefficient (R2) value is not met, the Cary will prompt you
with „Min R2 test failed‟. When you click „OK‟, the Cary will then prompt you with
„There is no valid calibration. Proceed in Abs (or Emission)?‟ If you click „Cancel‟,
the Concentration run will finish. If you click „Yes‟, the Cary will measure the
absorbance or emission of any presented samples, but will not generate a
concentration.
14 Measure sample concentration

a Once all the standards have been read, the ‘Present Sample’
dialog box will prompt you to place the appropriate sample
in the sample compartment. Click OK to measure the sample
and calculate its concentration. (If replicates have been
nominated, the concentration is calculated after the final
sample replicate is read.)
b Repeat for the remaining samples.
15 Save your data
a On the File menu, click Save Data As.
b Enter the ‘File name’ for this Concentration run.
16 Export your data
a On the File menu, click Export report (*.csv).
b Enter the ‘File name’ for this read.
c Click Save. The data will be stored as an ASCII spreadsheet,
with a *.csv file name extension.

How To…
Enzyme Kinetics
The following Enzyme Kinetics procedures are described:
 Performing a temperature-controlled run (Cary 50)
 Performing a temperature-controlled run using the Multicell
Holder (Cary 100–6000i)
Perform a temperature-controlled Enzyme Kinetics run (Cary 50)

This procedure describes how to perform a single cell, multi-rate
Enzyme Kinetics run at 37 °C using the single cell Peltier accessory
(Cary 50).
Setup dialog box
Click the Setup button or choose Setup from the Menu line to display
the ‘Setup’ dialog box and specify the method parameters for a new
method.
2 Ensure the Multicell Holder accessory is not selected
As various options do not become available until the appropriate
accessories are selected, you need to select these first.
Ensure ‘Use Cell Changer’ is not selected.
3 Set up accessories for reaction temperature control and
temperature display
a Under ‘Temperature’, select Automatic Temperature Setting
to enable the single cell Peltier accessory.
b Set the monitoring temperature by entering the Block
temperature as 37 °C.
c Under ‘Temperature Display’, select Probes 1 and 2 to view
the temperature of two temperature probes in the ‘Status
Display’ window.

How To…

a Enter the ‘Wavelength’ and ‘Ave. (averaging) Time’ you
require in the corresponding fields.
b Enter an upper and lower range value in the ‘Y min.’ and ‘Y
max.’ fields to specify the ordinate range.
5 Set up rate parameters
a Under ‘Collect Timing’, select Advanced Collect.
NOTE „Advanced Collect‟ enables you to collect data more frequently during the crucial
stages of your reaction, and to collect data less frequently where you know there
will be little activity.
b Enter the number of different reaction rates that you require

in the ‘Number of Stages’ field. The number you set here will
be reflected in the table below.
c Specify how long the Cary will wait after reading each cell
before it starts another reading cycle by setting the ‘Cycle
time’ for each rate stage.
d Specify the duration of the measurement run by setting the
‘Stop time’ for each rate stage.
6 Set up display options
Select ‘Individual Data’ to display the collected data of each sample
in individual graph boxes, or ‘Overlay Data’ to superimpose the
collected data of each sample in the run in one graph box.
7 Set up V0 calculation
Setup dialog box | Analyze page
a Set the start and stop times for your V0 calculation.
b Enter the correct product absorptivity and correct cell
pathlength for your reaction.

How To…
8 Set up calculations for the maximum rate (Vmax) and substrate

concentration that gives half the maximum rate (Km)
a Select the method by which the data obtained from your
selection under ‘Plot/Fit’ will be analyzed. Choose ‘Linear
Least Square’ or ‘Marquardt’.
b Choose the inhibitor model for your analysis. Select ‘Non
Competitive’, ‘Competitive’ or ‘Uncompetitive’.
c Select the Plot/Fit type/s that will be used to determine Vmax
and Km values.
d Select Auto Calculate to automatically perform Enzyme
Kinetics calculations on collected data at the end of each run.
These results will be displayed in the Report area.
c Set up your report style by selecting the appropriate items
under ‘Options’. For example, select ‘Auto Print’ to
automatically obtain a printout of your report. Select ‘Graph’
to include a graph in the generated report.
NOTE If „Auto Print‟ is selected, the system will send the report information to the
specified printer as well as to the Report area. However, if „Auto Print‟ is not
clicking „Report‟ on the „View‟ menu.
d Select the Autoconvert option you require. If you choose

‘Select for ASCII (csv)’ or ‘Select for ASCII (csv) with Log’, at
the end of the data collection, the system will automatically
generate a report and store the data in the Cary format as
well as ASCII XY pairs format in the current folder.

How To…

Select Storage On (Prompt at Start), to set the Cary to prompt you
for a file name before the start of an Enzyme Kinetics reaction.
View menu, to display information on your current reaction.
12 Finish setup
Click OK to save any changes you have made and close the ‘Setup’
dialog box.
a Click the Zero button or choose Zero from the Commands
menu. A ‘Loading Guide’ will be displayed.
b If you like, change the names of the blank sample.
c Place the blank solution in the correct cell position and click
OK. The system will perform an instrument zero on the blank
solution.
14 Start the run
a Click the Start button or choose Start from the Commands
menu. Do not add your active reagent at this time. The
system will display a ‘Loading Guide’.
b If you like, change the names of the sample.
c Place the sample solution in the correct cell position and
click OK. The system will set up the Graphics area and then
display the ‘Save As’ dialog box.
d Enter the ‘File name’ for this run and click Save. The ‘Sync
Start’ dialog box will be displayed.
e Add your active reagent just before the countdown reaches
0:00 or commence the data collection by clicking OK.
15 Enter substrate and inhibitor concentrations (two options)
Once the run has started:
a Open the ‘Setup’ dialog box by clicking the Setup button or
by choosing Setup from the Menu line.

How To…
b Click the Samples tab. In the ‘S’ column, enter the Substrate
concentration in micromoles. In the ‘I’ column, enter the
Inhibitor concentration in micromoles.
NOTE If no information has been entered during the collect, no calculation is

performed.
You can also use the ‘User Data Form’ to enter the Inhibitor and
Substrate concentrations:
a Open the ‘User Data Form’, by right-clicking in a graph box
and clicking User Data Form from the menu, or by choosing
User Data Form from the Graph menu.
b The table that appears has Data Names and may have V0
values already entered in the first two columns. In the third
and fourth columns, enter your values for [S] and [I] in
micromoles per liter.
c Click OK. Your [S] and [I] values are now ready to be used in
calculations, and the Cary will perform the calculations at
the end of the run.
Perform a temperature-controlled Enzyme Kinetics run using the

Multicell Holder (Cary 100–6000i)
This procedure describes how to perform a multicell, multi-rate
Enzyme Kinetics run at 37 °C using the Temperature Controller
accessory with the Multicell Holder accessory (Cary
100/300/4000/5000/6000i).
Setup dialog box
In Enzyme Kinetics, click the Setup button or select Setup from the

How To…
2 Set up the Multicell Holder accessory

a Select Use Cell Changer to enable the accessory.
b For Cary 100/300 instruments, choose the type of Multicell
Holder you are using (6 x 6 or 8 x 6). Ensure that you have
this accessory installed before starting the run.
NOTE If you are using a Series I 6 x 6 (Cary 100/300), you must calibrate the cell
changer using the Align application before starting experiments.
c Choose Select Cells and select the cells you require from the
available cells under ‘Use Cells’.
NOTE For Front Beam analysis, select „Cell 1–Cell 6‟ (6 x 6) or „Cell 1–Cell 8‟ (8 x 6).
This will ensure that all front cell positions in the Multicell Holder will be
measured during your enzyme kinetics analysis.
d Select Multi Zero to turn on the ‘Multi Zero’ facility.

e Ensure that Blank Correction is not selected.
temperature display
a If you are not using a Peltier-controlled accessory (for
example, the water-thermostatted 8 x 6), ensure that you
have the Temperature Controller accessory installed before
starting the run, and,
(i) Select Automatic Temperature Setting and select
Temperature Controller to enable the accessory.
(ii) Set the monitoring temperature by entering the Block
temperature as 37 °C. (The monitoring device is selected in
Step 4d).

How To…
b Under ‘Temperature Display’, select Block and Probe 1 to

view the temperature of the Multicell Holder block and one
temperature probe in the ‘Status Display’ window.
a Enter the ‘Wavelength’, ‘SBW’ (spectral bandwidth) and ‘Ave.
(averaging) Time’ you require in the corresponding fields.
b Select the ordinate mode you require. Click ‘Abs’ to specify
Absorbance mode or ‘%T’ to specify percent Transmittance.
NOTE The „%T‟ mode is used when performing fluorescence kinetics measurements
using the Total Fluorescence accessory or the Fluorescence Fiber Optic Probe.
c Enter an upper range and lower range value in the ‘Y min.’

d In the ‘Monitor’ field, choose where you are going to monitor
the temperature.
a Under ‘Collect Timing’, select Advanced Collect.
NOTE The „Advanced Collect‟ facility enables you to collect data more frequently during
the crucial stages of your reaction, and to collect data less frequently where you
know there will be little activity.

c Vary the number of data points collected per cell per run by
setting the ‘Dwell time’ for each rate stage.
d Specify how long the Cary will wait after reading each cell
before it starts another reading cycle, by setting the ‘Cycle

How To…
e Specify the duration of the measurement by setting the ‘Stop

6 Set up lamp and graphics options
the lamps at the end of a collect. This is especially useful
when performing data collections overnight or unattended
for long periods of time.
b Click UV/Vis if you want both lamps on during the run.
c Enter your required ‘Source Changeover’ and ‘Detector
Changeover’ (Cary 5000 only) wavelengths in the
corresponding fields. If you are using a Cary 5000/6000i, also
enter the detector and grating changeover wavelengths
(800 nanometers is recommended for both).
d Set the ‘Slit Height’ to full or reduced (Cary 4000/5000/6000i
only).
e Under ‘Display Options’, choose ‘Individual Data’ to display
the collected data of each sample in individual graph boxes.
Choose ‘Overlay Data’ to superimpose the collected data of
each sample in the Enzyme Kinetics run in one graph box.
7 Set up V0 calculation
a Set the start and stop times for your V0 calculation.
b Enter the required product absorptivity and cell pathlength
for your reaction.
8 Set up calculations for the maximum rate (Vmax) and substrate
concentration that gives half the maximum rate (Km)
a Under ‘Analyze’, select the method by which the data
obtained from your selection under ‘Plot/Fit’ will be
calculated. Choose ‘Linear Least Square’ or ‘Marquardt’.
b Choose the inhibitor model for your analysis.
c Select the Plot/Fit type/s that will be used to determine Vmax
and Km values.
d Select Auto Calculate to automatically perform enzyme
kinetics calculations at the end of each run.

How To…

specified printer as well as to the Report area.
d Select the Autoconvert option you require. If you choose

‘Select for ASCII (csv)’ or ‘Select for ASCII (csv) with Log’,
the system will automatically generate a report and store the
data in the Cary format as well as ASCII XY pairs format in
the current folder.
Select Storage On (Prompt at Start), to set up the Cary to prompt
you for a file name before the start of an Enzyme Kinetics reaction.
12 Finish setup
dialog box. Depending on the cells selected in the Multicell Holder,
the Cary may inform you that it will perform a dual single beam
calibration. Click OK.
a Click Zero or choose Zero from the Commands menu. A ‘Cell
Loading Guide’ will be displayed.

How To…
b If you like, change the names of the blank samples.

c Place the blank solution(s) in the correct cell positions and
click OK. The system will perform an instrument zero on the
blank solution(s).
NOTE If you had chosen not to use the „Multi Zero‟ facility in Step 2d, the system will
prompt you to enter the blank solution into the instrument. You must make sure
that you place the blank solution in the cell position that is currently in the light
path. Once you click „OK‟, the system will perform a zero on the cell position in
the light path, that is, the system will not reset the Multicell Holder to position 1.
14 Start the run

a Click Start or choose Start from the Commands menu. Do
not add your active reagent at this time. The system will
display a ‘Cell Loading Guide’.
b If you like, change the names of the samples.
c Place the sample solution(s) in the correct cell positions and
click OK. The system will set up the Graphics area and
display the ‘Save File’ dialog box.
d Enter the file name and click Save. The ‘Sync Start’ dialog
box will be displayed.
e Reset the Multicell Holder to position to cell 1 by clicking the
Reset Slide button. Add your active reagent just before the
countdown reaches 0:00, or commence the data collection by
clicking OK.
15 Enter substrate and inhibitor concentrations (two options)
Once the run has started:
a Open the ‘Setup’ dialog box by clicking the Setup button or
by choosing Setup from the Menu line.
b Click the Samples tab. In the ‘S’ column, enter the Substrate
concentration in micromoles. In the ‘I’ column, enter the
Inhibitor concentration in micromoles.

How To…
NOTE If no information has been entered during the collect, no calculation is

performed.
You can also use the ‘User Data Form’ to enter the Inhibitor and
Substrate concentrations:
a Open the ‘User Data Form’, by right-clicking in a graph box
and clicking User Data Form from the menu, or by choosing
User Data Form from the Graph menu.
b The table that appears has Data Names and may have V0
values already entered in the first two columns. In the third
and fourth columns, enter your values for [S] and [I] in
micromoles per liter.
c Click OK. Your [S] and [I] values are now ready to be used in
calculations, and the Cary will perform the calculations at
the end of the run.

How To…
Kinetics
The following Kinetics procedures are described:
 Performing a temperature-controlled Kinetics run (Cary 50)
 Performing a temperature-controlled Kinetics run using the
Multicell Holder (Cary 100–6000i)
 Performing a multi-wavelength, single cell, single rate Kinetics
run (Cary 100–6000i).
Perform a temperature-controlled Kinetics run (Cary 50)

This procedure describes how to perform a single cell, multi-rate
Kinetics run at 37 °C using the single cell Peltier accessory (Cary 50).
Setup dialog box
In Kinetics, click the Setup button or choose Setup from the Menu
line to display the ‘Setup’ dialog box and specify the method
2 Ensure the Multicell Holder accessory is not selected
Ensure ‘Use Cell Changer’ is not selected.
temperature display
a Under ‘Temperature’, select Automatic Temperature Setting
to enable the single cell Peltier accessory.
b Set the monitoring temperature by entering the block
temperature as 37 °C.
c Under ‘Temperature Display’, select Probe 1 to view the
temperature of one temperature probe in the ‘Status Display’
window.

How To…

a In the ‘Wavelength’ field, enter the wavelengths that you
would like to monitor.
b Enter an upper range and lower range value in the ‘Y min.’
c Select the abscissa (X) mode you require. Click ‘Min.’ to time
in minutes or ‘Sec.’ to time in seconds.
a Under ‘Collect Timing’, select Advanced Collect. This enables
you to set up different data collection procedures for the
multiple rates in your reaction.
NOTE „Advanced Collect‟ enables you to collect data more frequently during the crucial
stages of your kinetics reaction, and to collect data less frequently where you
know there will not be much activity.

c Set the ‘Cycle time’ for each rate stage to specify how long
the Cary will wait after reading each cell before it starts
another reading cycle.
d Set the ‘Stop time’ for each rate stage to specify the duration
of the measurement run.
6 Set up display options
Select ‘Individual Data’ to display a separate graph for each cell, or
‘Overlay Data’ to display all results in the one graph.
7 Set up analysis parameters
a Select Auto Calculate to automatically perform a rate
calculation on collected data at the end of each run.

How To…
b Select Advanced Calculate to set up multiple reaction rate

calculations for the Kinetics run.
c Enter the number of different rate calculations that you
require in the ‘Number of Stages’ field. The number you set
here will be reflected in the table below.
d Set up the stage ‘Start’ and ‘Stop’ times and select the
reaction order for each of these reaction stages.
NOTE If you select a „first order‟ or „second order‟ Simple Calculate rate calculation,
you can use the „Manual Guess‟ items to manually enter the parameters: A 0, Alnf
and Rate (k). It is presumed that you have a reasonable idea of the values for
these fit parameters, as they will be used as a first guess for the Marquardt non-
linear regression analysis.
If you do not choose ‘Manual Guess’ the Cary system will

automatically calculate the A0, Alnf and Rate values when the ‘Start’
button is clicked.
e Enter a value in the ‘Factor’ field to calculate enzyme
activity. This numerical multiplication factor is applied to the
absorbance.
f If you are performing a second order reaction, enter the
initial concentration of the substrate before reaction.
g Select Display Fit to automatically overlay the calculated
lines of best fit onto the plotted data.
boxes under ‘Options’. For example, select ‘Auto Print’ to

How To…
specified printer as well as the Report area. However, if „Auto Print‟ is not
d Select Include X-Y Pairs Table to view a list of abscissa

values and their corresponding ordinate values.
e Select the ‘Autoconvert’ option you require. If you select
‘ASCII’ or ‘ASCII with Log’, at the end of the data collection,
the current folder.
Select Storage On (Prompt at Start) to set the Cary to prompt you
for a file name before the start of a Kinetics run.
View menu, to display information on your current reaction.
11 Finish setup
a Click Zero, or choose Zero from the Commands menu. A
‘Loading Guide’ will be displayed.
b If you like, change the name of the blank.
c Place the blank solution in the sample compartment and click
solution.

How To…
13 Start the run

a Click the Start button to start a data collection. Alternatively,
choose Start from the Commands menu. Do not add your
active reagent at this time. The system will display the ‘Save
As’ dialog box.
b Enter the ‘File name’ for this Kinetics run and click Save.
The system will display a ‘Loading Guide’.
c If you like, change the name of the sample.
d Place the sample solution in the correct position and click
OK. The system will set up the Graphics area and then
display the ‘Sync Start’ dialog box.
e Add your active reagent just before the countdown reaches
0:00, or commence the data collection by clicking OK.
Perform a temperature-controlled Kinetics run using the Multicell

This procedure describes how to perform a multicell, multi-rate
Kinetics run at 37 °C using the Temperature Controller accessory
with the Multicell Holder accessory (Cary 100/300/4000/5000/6000i).
Setup dialog box
method.

How To…
c Click Select Cells and select the cells you require from the
For Front Beam analysis, select ‘Cell 1 to Cell 6’ (6 x 6) or
‘Cell 1 to Cell 8’ (8 x 6). This will ensure that all front cell
positions in the Multicell Holder will be measured during
your Kinetics analysis.
d Select Multi Zero.
e Ensure ‘Blank Correction’ is not selected.
temperature display
(ii) Set the monitoring temperature by entering the Block
Step 4e).
turn on monitoring of the Multicell Holder block and one
temperature probe.
a Enter the ‘Wavelength’, ‘SBW’ (spectral bandwidth) and ‘Ave.
(averaging) Time’ you require in the corresponding fields.
b Select the ordinate mode you require. Click ‘Abs’ to specify
absorbance mode or ‘%T’ to specify percent transmittance.

How To…
c Enter an upper range and lower range value in the ‘Y min.’

d Select ‘Min.’ or ‘Sec.’ to set the abscissa (X).
e In the ‘Monitor’ field, choose where you are going to monitor
the temperature. The ‘Start’ button will not be enabled until
the temperature of the selected monitor is within 0.5 °C of
the block temperature set on the ‘Accessories’ page (Step 3a
(ii)).
know there will be little activity.

c Vary the number of data points collected per cell, per run, by
setting the ‘Dwell time’ for each rate stage.
d Specify how long the Cary will wait after reading each cell
e Specify the duration of the Kinetics run by setting the ‘Stop
when performing Kinetics data collections overnight or
b Click the UV/Vis button if you want both of the lamps on
during the run.

How To…

enter the detector and grating changeover wavelengths (800
nanometers is recommended for both).
only).
e Under ‘Display Options’ choose ‘Individual Data’ to display
each sample in the Kinetics run in one graph box.
a Select Auto Calculate to automatically perform a rate
calculation at the end of each run.
b Select Advanced Calculate to set up multiple rate
calculations for the Kinetics run.
c Enter the number of different rate calculations that you
require in the ‘Number of Stages’ field. The number you set
here will be reflected in the table below.
d Set up the stage ‘Start’ and ‘Stop’ times and select the
reaction order for each of these reaction stages.
NOTE If you select a „first order‟ or „second order‟ Simple Calculate rate calculation,
you can use the items under „Manual Guess‟ to manually enter the parameters:
A0, Alnf and Rate (k).
e Enter a value in the ‘Factor’ field to calculate enzyme

activity. This multiplication factor is applied to the
absorbance.
f If you are performing a second order reaction, enter the
initial concentration of substrate before reaction.
g Select Display Fit to automatically overlay the calculated

How To…


the current folder.
Select Storage On (Prompt at Start) to set the Cary to prompt you
for a file name, before the start of a Kinetics reaction.
11 Finish setup
dialog box.
a Click Zero or from the Commands menu, choose Zero. A
‘Cell Loading Guide’ will be displayed.

How To…
b If you like, change the names of the blank samples.

c Place the blank solution(s) in the correct cell positions and
blank solution(s).
13 Start the run
a Click the Start button or from the Commands menu, choose
Start. Do not add your active reagent at this time. The
system will display the ‘Save As’ dialog box.
b Enter the ‘File name’ of this Kinetics run and click Save. The
system will display a ‘Cell Loading Guide’.
c If you like, change the names of the samples.
d Place the sample solution(s) in the correct cell positions and
click OK. The system will set up the Graphics area and then
the ‘Sync Start’ dialog box will be displayed.
e Reset the Multicell Holder position to cell 1 by clicking the
Reset Slide button.
f Add your active reagent just before the countdown reaches
Perform a multi-wavelength, single cell, single rate Kinetics run (Cary

100–6000i)
This procedure describes how to perform a multi-wavelength, single
cell Kinetics run at ambient temperature (Cary
100/300/4000/5000/6000i).
Setup dialog box
In Kinetics, click the Setup button or choose Setup from the Menu
a In the ‘Wavelength’ field, enter the relevant wavelengths.
b In the ‘SBW’ field, enter the required spectral bandwidth.

How To…
c In the ‘Ave. (averaging) Time’ field, enter the required value.

d Select the ordinate mode you require. Click ‘Abs’ to specify
the absorbance mode or ‘%T’ to specify percent
transmittance.
e Enter an upper range and lower range value in the ‘Y min.’
and ‘Y max.’ fields to specify the ordinate range.
f Select ‘Min.’ or ‘Sec.’ to set the abscissa (X).
a Under ‘Collect Timing’, select Simple Collect to set up a
single rate for your reaction.
b Specify how long the Cary will wait after reading each cell
before it starts another reading cycle by setting the cycle
time.
c Specify the duration of the Kinetics run by setting the stop
time.
when performing Kinetics data collections overnight or
b Click the UV/Vis button if you want both lamps on during the
run.
only).
e Under ‘Display Options’, select ‘Individual Data’ to display
Select ‘Overlay Data’ to superimpose the collected data of
each sample in the Kinetics run in one graph box.

How To…
5 Set up accessories
Click the Accessories tab and ensure that no accessories are
selected.
a Select ‘Auto Calculate’ to automatically perform a rate
calculation at the end of each run. Or select ‘Simple
Calculate’ to perform one rate calculation for the entire
Kinetics run.
b Set up the reaction start and stop times and select the
reaction order.
c If you select a ‘first order' or ‘second order' Simple Calculate
rate calculation, you can use the ‘Manual Guess’ items to
manually enter the parameters: A0, AInf and Rate (k).
NOTE If you do not choose „Manual Guess‟, the Cary system will automatically
calculate the values: A0, AInf and Rate when the „Start‟ button is clicked.
d Enter a value in the ‘Factor’ field to calculate enzyme

activity. This multiplication factor is applied to the
absorbance.
e If you are performing a second order reaction, enter the
initial concentration of substrate before reaction.
f Select Display Fit to automatically overlay the calculated

How To…

specified printer as well as the Report area.

e Select the autoconvert option you require. If you select
the current folder.
Select Storage Off. The method, collected data and report will not be
saved. Or, select ‘Storage On Prompt at Start’ to set the Cary to
prompt you for a file name before the start of a run; or ‘Storage On
Prompt at End’ to set the Cary to prompt you for a file name at the
end of a run.
10 Finish setup
a Click Zero. Alternatively, choose Zero from the Commands
menu. A ‘Loading Guide’ will be displayed.

How To…
c Place the blank solution in the sample compartment and click

solution(s).
12 Start the run
a Click the Start button, or choose Start from the Commands
menu. Do not add your active reagent at this time. The
system will display a ‘Loading Guide’.
b If you like, change the names of the sample.
c Place the sample solution in the front cell holder and click
OK. The system will set up the Graphics area and then
display the ‘Sync Start’ dialog box.
d Add your active reagent just before the countdown reaches
0:00, or commence the data collection by clicking OK. The
Cary will start the Kinetics data collection.
13 Save your data
a On the File menu, and click Save Data As.
b Enter the ‘File name’ for this Kinetics run.

How To…
RNA-DNA Estimation
The following RNA-DNA procedures are described:
 Performing a run using the Multicell Holder (Cary 50)
 Performing a temperature-controlled run using the Multicell
Perform an RNA-DNA run using the Multicell Holder (Cary 50)

This procedure describes how to perform a multicell wavelength scan
with baseline correction, multi zero and multi baseline (Cary 50).
Setup dialog box
method.
Setup dialog box | Cary and Baseline pages
a Enter the first and second wavelengths at which you would
like to measure your sample.
b If you require background correction, select Background
Correction and enter a background wavelength in the
corresponding field.
c If you are performing a wavelength scan, select Scan
Samples. The ‘Display Options’ will be activated and the
‘Baseline’ page will be displayed.
d Enter a ‘Start’ and ‘Stop’ wavelength value and select a ‘Scan
Rate’.
e Under ‘Display Options’, select the way in which you want
the data displayed as it is collected.
Choose ‘Individual Data’ to display the collected data of each
sample in individual graph boxes. Choose ‘Overlay Data’ to
superimpose the collected data of each sample in the run in
one graph box.

How To…
f Click the Baseline tab and select Baseline correction.

g To use a previously stored baseline, click the Baseline button
and browse for the appropriate file. Otherwise, you can
perform your baseline correction at the beginning of the run.
3 Set up Multicell Holder accessory
Setup dialog box | Accessories 1 page
a Select Use Cell Changer to enable the accessory. Ensure that
you have this accessory installed before starting the run.
b Click Select Cells and select the cells you require from the
c Select Multi Zero to turn on the ‘Multi Zero’ facility.
d Select Multi Baseline to turn on the ‘Multi Baseline’ facility.
a Enter the number of samples that you are going to use in the
‘Number of Samples’ field. The ‘Sample Names’ list below will
expand or contract to match your choice.
b In the ‘Sample Names’ list, enter the name of each sample.
You can enter up to 20 characters for each name.
click the ‘Increment’ button.
Click the ‘Import’ button to use sample names stored in a text
file.
c Clear the red tick in the first column for any samples that
you do not wish to analyze.
d If you would like multiple readings of the same aliquot, select
Replicates and enter the number of replicates required in the
field that appears.
If you would like to calculate any Warburg Christian or 260 nm
Factor parameters, make your selections here.

How To…

automatically obtain a printout of your report. If you have
selected ‘Scan Samples’ on the ‘Cary’ page, select ‘Graph’ to
include a graph in the generated report.
d Select the ‘Autoconvert’ option you require. If you choose

the end of the data collection the system will automatically
for a file name before the start of the RNA-DNA run.
Select Show Status Display on any of the Setup pages or choose
Status Display from the View menu to display information about
your current reaction.
9 Finish setup
dialog box.

How To…

b Place the blank solution(s) in the correct cell positions and
blank solution(s).
NOTE If you have chosen not to use the „Multi Zero‟ facility in Step 3c, the system will
prompt you to enter one blank solution into the instrument. In this case, it is
important to ensure that you place this blank solution in the cell position that is
currently in the light path. On clicking „OK‟, the system will perform a zero on the
cell position in the light path, that is, the system will not reset the Multicell
Holder to position 1.
11 Collect baselines (if not using a stored baseline)

If you have selected to perform a baseline correction (Step 2f) and
are not using a stored baseline, take a baseline reading now by
following the steps below. Otherwise, proceed to Step 12.
a Click Baseline to collect a baseline for each cell. A ‘Cell
b Load the blank/s as depicted.
c Click OK. On completion of the baseline collections, the word
‘baseline’ will be displayed in red above the ordinate
instrument status reading.
12 Start the run
menu to start a data collection. The system will display the
‘Save As’ dialog box.
b Enter the ‘File name’ for this RNA-DNA run in the ‘File name’
field and click Save. The system will display a ‘Cell Loading
Guide’.
click OK. The system will then begin reading the samples and
printing the results to the Report area.

How To…
Perform a temperature-controlled RNA-DNA run using the Multicell

This procedure describes how to perform a multicell RNA-DNA run
with multi zero and multi baseline at 37 °C using the Temperature
Controller accessory with the Multicell Holder accessory (Cary
100/300/4000/5000/6000i).
Setup dialog box
In RNA-DNA, click the Setup button or choose Setup from the Menu
c Click Select Cells and select the cells you require under ‘Use
Cells’. For double beam analysis, select ‘Cell 1 through to the
Cell 6’ (6 x 6) or ‘Cell 1 through to Cell 8’ (8 x 6).
d Select Multi Zero.
The ‘Multi Baseline’ facility will not be accessible until completing
Step 4.

How To…

temperature display
(ii) Set the monitoring temperature by entering the block
Step 4).
Setup dialog box | Cary and Baseline pages
a Enter the first and second wavelengths at which you would
like to measure your sample/s.
b If required, select Background Correction and enter a
background wavelength in the corresponding field.
c Choose the desired temperature monitoring device in the
‘Monitor’ field. The ‘Start’ button will not be enabled until the
temperature of the selected Monitor is within 0.5 °C of the
Block temperature set on the ‘Accessories 1’ page (Step 3a
(ii)).
d To perform a wavelength scan, select Scan Samples and set
the ‘Start’, ‘Stop’ and ‘Rate’ for the scan. The ‘Baseline’ tab
will be displayed.
e Click the Baseline tab.
f Select Baseline correction. To use a previously stored
baseline, click the ‘Baseline’ button and browse for the
appropriate file. Otherwise, you can perform your baseline
correction at the beginning of the run. To perform a ‘Multi
Baseline’, you can now go back to the ‘Accessories 1’ page
and select ‘Multi Baseline’.

How To…
5 Set up source, SBW and display options

d Enter the spectral bandwidth of your instrument in the ‘SBW’
field.
e Set the appropriate ‘Slit Height’ (Cary 4000/5000/6000i only).
f Under ‘Display Options’, select ‘Individual Data’ to display
each sample in the run in one graph box.
a Enter the number of samples that you are going to use in the
‘Number of Samples’ field. The ‘Sample Names’ list below will
expand or contract to match your choice.
b In the ‘Sample Names’ list, enter the name of each sample.
You can enter up to 20 characters for each name.
Click the ‘Import Names’ button to use names from a stored
text file.
c Clear the red tick for any samples not required.
d If you would like multiple readings of the same aliquot, select
Replicates and enter the number of replicates required.

How To…

If you would like to calculate any Warburg Christian coefficients or
260 nm Factor parameters, make your selections here.
automatically obtain a printout of your report. If you have
selected ‘Scan Samples’ on the ‘Cary’ page, select ‘Graph’ to
include a graph in the generated report.
d Select the ‘Autoconvert’ option you require. If you choose

the end of the data collection, the system will automatically
for a file name before the start of the RNA-DNA run.
Set up visual system monitoring

How To…
10 Finish setup
dialog box.
b Place the blank solution(s) in the correct cell positions and
blank solution(s).
NOTE If you have chosen not to use the „Multi Zero‟ facility in Step 2d, the system will
prompt you to enter one blank solution into the instrument. In this case, it is
important to ensure that you place this blank solution in the cell position that is
currently in the light path. On clicking „OK‟, the system will perform a zero on the
cell position in the light path, that is, the system will not reset the Multicell
Holder to position 1.
12 Perform a baseline correction (if not using a stored baseline)

If you have selected to perform a baseline correction (Step 4f) and
are not using a stored baseline, take a baseline reading now by
following the steps below. Otherwise, proceed to Step 13.
a Click Baseline. A ‘Cell Loading Guide’ will be displayed.
b Load the blank/s as shown.
c Click OK. The system will collect a baseline for each cell in
use and the word ‘baseline’ will be displayed above the
ordinate instrument status reading.
13 Start the run
a Click the Start button to start a data collection. Alternatively
you can choose Start from the Commands menu. The ‘Save
As’ dialog box will be displayed.
b Enter the ‘File name’ for this RNA-DNA run and click Save.
The system will display a ‘Cell Loading Guide’.

How To…

click OK. The system will then begin reading the samples and
printing the results to the Report area.
Scan
The following Scan procedures are described.
 Performing a scan with baseline correction (Cary 50)
 Performing a scan with baseline correction (Cary 100–6000i)
 Performing a scan in Signal-to-Noise mode (Cary 100-6000i)
 Performing a scan in Independent mode (Cary 5000/6000i)
Perform a scan with baseline correction (Cary 50)

This procedure describes how to perform a wavelength scan from
800 nm to 200 nm with baseline correction using a Cary 50.
Setup dialog box
In Scan, click Setup or choose Setup from the Menu line to display
method.
a Set the wavelength range for the scan by entering the values
you require in the ‘Start’ and ‘Stop’ fields.
b In the ‘Y Mode’ field, select the ordinate mode in which you
want the collect data to be displayed.
c Enter an upper and lower range value in the ‘Y min.’ and ‘Y
max.’ fields to specify the displayed ordinate range.
d Make sure that ‘Cycle Mode’ is not selected.
e Set the ‘Beam Mode’ for the run. This should be set to Dual
Beam.

How To…
f Under ‘Scan Controls’, select Simple and click a scan speed

button. Alternatively, you can select ‘Advanced’ and enter an
‘Ave. Time’ and ‘Data Interval’ (the Cary will then select the
Scan Speed).
g Under ‘Display Options’, select the way in which you want
the data displayed as it is collected. Choose ‘Individual Data’
to display the collected data of each sample in individual
graph boxes. Choose ‘Overlay Data’ to superimpose the
collected data of each sample in the Scan run in one graph
box.
3 Set up the baseline correction
Setup dialog box | Baseline page
Select Baseline Correction. This will force the Cary to perform a
baseline correction on the sample data. The correction will be
performed on each point before it is displayed.
NOTE You can use a stored baseline. To do this, click „Baseline‟ and open the saved
*.csw baseline file.

Make sure that no options are selected on this page and that no
accessories are installed.

How To…

‘Parameters’ to include your method parameters in the
report. Select ‘Graph’ to include a graph in the generated
report.
specified printer as well as the Report area. However, if „AutoPrint‟ is not
d Set up the ‘Peak Table’ reporting.

(i) Select Peak Labels.
(ii) Click the Peak Information button and choose the type of
‘Peak Labels’, the ‘Peak Style’ and set the ‘Peak Threshold’.
Click OK.
(iii) Select Maximum Peak to report the peak with the largest
peak threshold that exceeds the Peak Threshold value.
(iv) Select All Peaks to report all peaks meeting the Peak
Style criterion and exceeding the Threshold value.
e Set up ‘X-Y pairs’ reporting, if required. You can use the
actual Data Interval by which the data was collected or you
can make the Cary interpolate the points to a new Interval.
f Select the ‘Autoconvert’ option you require. If you select
Select Storage On (Prompt at Start).

How To…

Select Show Status Display on any Setup page or from the View
menu to display information about your current reaction.
8 Finish setup
Click Zero or choose Zero from the Commands menu to zero the
system.
10 Measure a baseline
a Click Baseline or choose Baseline from the Commands
menu.
b When prompted, insert the blank sample into the sample
compartment front beam and click OK.
The Cary will collect the baseline scan. After the collection, the word
‘baseline’ will be displayed in red in the ordinate status box,
indicating that you are in baseline correction mode and you have a
valid baseline file for the correction.
NOTE If the word ‟baseline‟ is gray and in italics, the baseline file is still valid. The gray
and italics indicate that the Cary is idling outside the abscissa range of the
baseline file.
11 Start the run

Click the Start button to start a data collection. Alternatively, you
can choose Start from the Commands menu.
12 Specify a file name for the data and sample names
a When you click Start, the ‘Save As’ dialog box will be
displayed. Enter the appropriate name for your Scan run and
click Save.

How To…
b The ‘Sample Name’ dialog box will be displayed. Enter the

appropriate name for your sample and click OK. The Scan
run will commence and the corrected trace will be displayed
in the Graphics area. At the end of the run, the Cary will
create the report and also print it if ‘AutoPrint’ was selected
on the ‘Reports’ page of the ‘Setup’ dialog box.
Perform a scan with baseline correction (Cary 100–6000i)

800 nm to 200 nm with baseline correction (Cary
100/300/4000/5000/6000i).
Setup dialog box
In Scan, click the Setup button or choose Setup from the Menu line
to display the ‘Setup’ dialog box and specify the method parameters
for a new method.
a Set the appropriate abscissa mode in the ‘X Mode’ field.
b Set the wavelength range by entering the values you require
in the ‘Start’/‘Stop’ fields.
c In the ‘Y Mode’ field, select the ordinate mode.
d Enter an upper and lower range in the ‘Y min.’ and ‘Y max.’
fields to specify the displayed ordinate range.
e Set the speed of the data collection by setting the ‘Ave. Time’
and ‘Data Interval’. The Data Interval is the wavelength
increment between data points. The ‘Scan Rate’ will
automatically update when selected.
f Make sure ‘Cycle Mode’ is not selected.
3 Set up SBW, lamp and graphics options
a Set the ‘SBW’ (spectral bandwidth).
b Set the ‘Beam Mode’ for the run (usually ‘Double’).

How To…
c For Cary 5000/6000i, the fixed SBW is used to automatically

alter the Energy level and maintain a constant signal level.
d For Cary 4000/5000/6000i, set the ‘Slit Height’ to Full.
e Click the UV/Vis button if you want both lamps on during the
run.
f Select Auto Lamps Off if you want to automatically turn off
when performing Scan runs overnight, or unattended for long
periods of time.
g Do not select ‘Signal-to-Noise Mode’.
h Under ‘Display Options’, select ‘Individual Data’ to display
each sample in the Scan run in one graph box.
Setup dialog box | Independent page
If you are using a Cary 5000/6000i, do not change any options on the
‘Independent’ page.
Select Baseline Correction to perform a baseline correction on each
sample data point.
NOTE You can use a stored baseline by clicking „Baseline‟ and opening the saved *.csw
baseline file.


How To…

b Enter a comment relating to your sample in the ‘Comment’
field.
to include a graph in the printed report.

(ii) Click the Peak Information button and choose the ‘Peak
Type’, the ‘Labels Type’, and set the ‘Peak Threshold’. Click
OK.
actual Data Interval by which the data was collected, or you

How To…

Select Storage On (Prompt at Start).
Set up visual system monitoring
Select Show Status Display on any Setup page, or from the View
menu, to display information about your current reaction.
8 Finish setup
Click Zero or choose Zero from the Commands menu to perform a
zero.
menu.
compartment front beam and click OK. After the baseline is
collected, the word ‘baseline’ will be displayed in red in the
ordinate status box, indicating that you are in baseline
correction mode and have a valid baseline file for the
correction.
baseline file.
11 Start the run

Click the Start button or choose Start from the Commands menu to
start the Scan run.

How To…
12 Name the data file and samples

a When you click Start, the ‘Save As’ dialog box will be
displayed. Enter the appropriate ‘File name’ for the data and
click Save.
b The ‘Sample Name’ dialog box will be displayed. Enter the
appropriate name for your sample and click OK. The Scan
run will commence and the corrected trace will be displayed
in the Graphics area. At the end of the run, the Cary will
create the report and also print it, if ‘AutoPrint’ has been
selected on the ‘Reports’ page of the ‘Setup’ dialog box.
Perform a scan in Signal-To-Noise mode (Cary 100-6000i)

800 nm to 200 nm in signal-to-noise mode
(Cary 100/300/4000/5000/6000i).
Setup dialog box
for a new method.
a Set the appropriate abscissa mode for the scan in the ‘X
Mode’ field.
b Set the wavelength range for the scan by entering the values
fields to specify the displayed ordinate range. You can click
the ‘Autoscale’ button during the data collection to
automatically scale the display.

How To…
f Make sure that ‘Cycle Mode’ is not selected.

a Set the ‘SBW’ (spectral bandwidth).
b If you are using a Cary 100/300/5000/6000i, set the ‘SBW’ for
the run. A good starting point is 2 nanometers if your method
does not specify a SBW.
c Set the ‘Beam Mode’ for the run (usually ‘Double’).
d For Cary 5000/6000i, the fixed SBW is used to automatically
alter the Energy level and maintain a constant signal level.
e For Cary 4000/5000/6000i, set the ‘Slit Height’ to Full.
f Click the UV/Vis button if you want both the lamps on during
the run.
g Select Auto Lamps Off if you want to automatically turn off
the lamps at the end of a collect. This option is especially
useful when performing Scan runs overnight or unattended
h Check Signal-to-Noise Mode, and set the required values for
the Acceptable S/N and the S/N Timeout.
i Under ‘Display Options’, select ‘Individual Data’ to display
Choose the ‘Overlay Data’ to superimpose the collected data
of each sample in the Scan run in one graph box.
On the ‘Independent’ page, make sure ‘Independent Control’ is not
selected.
Select ‘Baseline Correction’ to perform a baseline correction on each
sample data point.

How To…
baseline file.



How To…
Type’, the ‘Labels Type’, the and set the ‘Peak Threshold’.
Click OK.
Select Storage On (Prompt at End).
9 Finish setup
Click Zero, or choose Zero from the Commands menu to perform a
zero.
menu.

How To…

correction.
NOTE If the word „baseline‟ is gray and in italics, the baseline file is still valid. The gray
baseline file.
12 Start the run

Click the Start button or choose Start from the Commands menu.
13 Name your samples
Once you click ‘Start’, the ‘Sample Name’ dialog box will be
displayed. Enter the appropriate name for you sample and click OK.
The Scan run will commence and the corrected trace will be
displayed in the Graphics area.
If for a particular point, the set Acceptable S/N cannot be met in the
set ‘S/N Timeout’ time, the Cary will collect the point as normal
(using the S/N Timeout as the Ave. Time) and display the message
SNR Timeout in the hardware status area.
14 Save your data
When the Cary has measured the sample, the ‘Save As’ dialog box
will be displayed. Enter the appropriate name for your sample and
click Save. The Cary will then create the report and print it, if
‘AutoPrint’ has been selected on the ‘Reports’ page of the ‘Setup’
dialog box.

How To…
Perform a scan in Independent mode (Cary 5000)

2200 nm to 400 nm in Independent mode (Cary 5000 only).
Setup dialog box
for a new method.
a Set the appropriate abscissa mode for the scan in the ‘X
Mode’ field.
fields to specify the displayed ordinate range. You can use
the ‘Autoscale’ button during the data collection to
automatically scale the display.
e Do not set any Scan Controls, such as ‘Ave. Time’ or ‘Data
Interval’, as the settings in Independent mode will override
these.
f Make sure that ‘Cycle Mode’ is not selected.
a Do not set the ‘SBW’ (spectral bandwidth) or Energy, as the
settings in Independent mode will override these.
b Set the ‘Beam Mode’ for the run (usually ‘Double’).
c Set the ‘Slit Height’ to Full.
d Click the UV/Vis button if you want both lamps on during the
run.

How To…
e Select Auto Lamps Off if you want to automatically turn off

the lamps at the end of a collect. This option is especially
useful when performing Scan runs overnight or unattended
f Under ‘Display Options’, select ‘Individual Data’ to display
each sample in the Scan run in one graph box.
4 Set up the signal level controls (SBW/energy) for the scan
a Select Independent Control.
b Select the Auto ‘Measurement Mode’. This will mean that the
Cary will use a Fixed SBW (spectral bandwidth) in the UV-Vis
region and a fixed Energy level in the near infra-red region.
c Under ‘UV-Vis’, set the parameters you require in the UV-Vis
region. Good starting values are: Ave. Time = 0.1, Data
Interval = 1, Scan Rate = 600, SBW = 2.
d Under ‘NIR’, set the parameters you require in the near infra-
red region. Good starting values are: Ave. Time = 0.1, Data
Interval = 4, Scan Rate = 2400, Energy=1.
5 Set up baseline correction
Select Baseline Correction to perform a baseline correction on each
sample data point.
baseline file.


How To…


Type’, the ‘Labels Type’, and set the ‘Peak Threshold’. Click
OK.

How To…

10 Finish setup
Click Zero to zero the system. Alternatively, choose Zero from the
Commands menu to perform a zero.
menu.
correction.
baseline file.
13 Start the run

Click the Start button or choose Start from the Commands menu.

How To…
14 Set up sample names

When you click Start, the ‘Sample Name’ dialog box will be displayed.
Enter the appropriate name for you sample and click OK. The Scan
run will commence and the corrected trace will be displayed in the
Graphics area.
15 Save your data
When the Cary has measured the sample, the ‘Save As’ dialog box
will be displayed. Enter the appropriate name for your sample and
click Save. The Cary will then create the report and print it, if
‘AutoPrint’ was selected on the ‘Reports’ page of the ‘Setup’ dialog
box.
Scanning Kinetics
The following Scanning Kinetics procedures are described:
 Collecting data (Cary 50)
 Collecting data using the Multicell Holder with temperature
control (Cary 100–6000i)
Collect data (Cary 50)

This procedure describes how to perform a multicell, multi-stage
wavelength scan with baseline correction from 600 to 500 nm at
ambient temperature (Cary 50). You can then use the data to create
kinetics continuums in order to calculate reaction rates.
Setup dialog box
In Scanning Kinetics, click Setup or choose Setup from the Menu line
for a new method.
a Set the wavelength range for the scan by entering the values

How To…
b Enter an upper and lower range value in the ‘Y min.’ and ‘Y

max.’ fields to specify the displayed ordinate range.
c You now need to set the speed of the data collection by
setting the ‘Ave. Time’ and ‘Data Interval’. In the ‘Ave.
(averaging) Time’ field, enter the required value. 0.1 seconds
is a good starting value.
d In the ‘Data Interval’ field, enter the wavelength increment
you require between data points. 0.5 nanometers is a good
starting point. The Cary will automatically update the ‘Scan
Rate’ field when you select it.
the crucial stages of your reaction, and less frequently where you know there will
not be much activity.

c Specify how long the Cary will wait after reading each cell
d Specify the duration of the Scanning Kinetics run by setting
the ‘Stop time’ for each rate stage.
4 Select the baseline correction type
Select Baseline correction. This will force the Cary to use a baseline
scan to perform a baseline correction on the sample data. The
correction will be performed on each point before it is displayed.

How To…
NOTE You can use a stored baseline. To do this, click the „Retrieve Baseline file‟ button
and open the saved *.csk baseline file.

a Ensure that you have the appropriate accessories installed
before starting the run.
b Select Use Cell Changer to enable the Multicell Holder
accessory.
c Click Select Cells and select the cells you require from the
This page is used for post-run analysis.
‘Parameters’ to include your experimental parameters in the
report. Select ‘Graph’ to include a graph in the generated
report.

How To…

‘ASCII’ or ‘ASCII with Log’, at the end of the data collection
the current folder.
for a file name before the start of the Scanning Kinetics run.
10 Finish setup
dialog box.
If you do not have a valid baseline file, the Cary will prompt you to
click Baseline.
a Click Baseline to set up the baseline collection.
c Insert blank samples into the cell changer to collect the
0Abs/100%T baseline scans, and click OK.

How To…
The system will set up the Graphics area and the Cary will collect the
baseline scan. After the collection, the word ‘baseline’ will be
displayed in red in the ordinate status box, indicating that you are in
baseline correction mode and you have a valid baseline file for the
correction.
a Click Zero or choose Zero from the Commands menu to
perform a zero.
b If you like, change the name of the blank/s.
c Insert blank samples into the appropriate positions of the
Multicell Holder and click OK.
13 Start the run
a Click Start or choose Start from the Commands menu to
start a data collection. Do not add your active reagent at
this time.
NOTE At this point, the system will display the „Save File‟ dialog box if you have
selected „Storage On (Prompt at Start)‟ on the „Auto Store‟ page of the „Setup‟
dialog box. If so, enter the file name for this Scanning Kinetics run in the „File
name‟ field and click Save.
b The system will display a ‘Cell Loading Guide’. If you like,

change the names of the samples.
click OK. The ‘Sync Start’ dialog box will be displayed.
At the end of the run, you can determine the actual Stop Time by
observing the last value in the ‘Time’ column of the ‘User Data Form.’

How To…
Collect data using the Multicell Holder with temperature control

(Cary 100–6000i)
This procedure demonstrates how to perform a multicell, multi-stage
wavelength scan with baseline correction from 600 to 500 nm at
37 °C using the Temperature Controller accessory with the Multicell
Holder accessory (Cary 100/300/4000/5000/6000i). You can then use
the data to create kinetics continuums in order to calculate reaction
rates.
Setup dialog box
In Scanning Kinetics, click the Setup button or choose Setup from
the Menu line to display the ‘Setup’ dialog box and specify the
method parameters for a new method.
2 Select the baseline correction type
Select Baseline Correction. This will force the Cary to use a baseline
scan to perform a baseline correction on the sample data. The
correction will be performed on each point before it is displayed.
NOTE You can use a stored baseline. To do this, click the „Retrieve Baseline File‟ button
and open the saved *.csk baseline file.

a Ensure that you have the appropriate accessories installed
before starting the run.
b Select Use Cell Changer to enable the Multicell Holder
accessory.
c Choose the type of Multicell Holder you are using (6 x 6 or
8 x 6).

How To…
NOTE If you are using a Series I 6 x 6x (Cary 100/300), you must calibrate the cell
d Click Select Cells and select the cells you require under ‘Use
Cells’.
For Front Beam analysis, select ‘Cell 1–Cell 6’ (6 x 6) or
‘Cell 1–Cell 8’ (8 x 6). This will ensure that all front cell
positions in the Multicell Holder will be measured during
your Scanning Kinetics analysis.
e Select Multi Zero.
temperature display
starting the run.
b Select Automatic Temperature Setting and click
c Set the monitoring temperature by entering the block
Step 6d.)
d Under ‘Temperature Display’, select Block and Probe 1 to
a Set the appropriate abscissa mode for the scan in the
‘X Mode’ field.
c In the ‘Y Mode’ field, select the ordinate mode you require.
d Enter an upper and lower range value for ‘Y min.’ and
‘Y max.’ to specify the displayed ordinate range.

How To…
Under ‘Collect Timing’, select Advanced Collect to set up different
data collection procedures for the multiple rates in your reaction.
know there will not be much activity.
a Enter the number of different reaction rates that you require

b Specify how long the Cary will wait after reading each cell
c Specify the duration of the Scanning Kinetics run by setting
the ‘Stop time’ for each rate stage.
d Choose the desired temperature monitoring device in the
‘Monitor’ field. The ‘Start’ button will not be enabled until the
temperature of the selected monitor is within 0.5 °C of the
block temperature set on the ‘Accessories’ page (Step 4c).
7 Set up spectral bandwidth or energy and lamp options
a Set the ‘SBW’ (spectral bandwidth) for the run.
b Set the ‘Slit Height’ to Full.
c Select Auto Lamps Off if you want to automatically turn off
d Click the UV/Vis button if you want both lamps on during the
run.

How To…
e Enter your required ‘Source Changeover’ wavelength in the

corresponding field. If you are using a Cary 5000/6000i, also
This page is used for post-run calculations.

e Select the ‘Autoconvert’ option you require.
f If you choose ‘Select for ASCII (csv)’ or ‘Select for ASCII
(csv) with Log’, at the end of the data collection the system
will automatically generate a report and store the data in the
Cary format as well as ASCII XY pairs format in the current
folder.
for a file name before the start of the Scanning Kinetics run.

How To…

12 Finish setup
dialog box. Depending on the cells selected in the Multicell Holder,
the Cary may then inform you that it will perform a dual single beam
calibration. Click OK.
a If you do not have a valid baseline file, the Cary will prompt
you to click Baseline to set up the baseline collection.
change the name of the blank.
c Insert blank samples into the cell changer to collect the
0Abs/100%T baseline scans and click OK.
The system will set up the Graphics area and the Cary will collect the
baseline scan. After the collection, the word ‘baseline’ will be
displayed in red in the ordinate status box, indicating that you are in
baseline correction mode and you have a valid baseline file for the
correction.
If you want to use the baseline again with other samples, save the
method using ‘Save Method As’ on the ‘File’ menu. Then, when you
re-open the method, the baseline will also open and be ready to use.
It is preferable to save the baseline with the method, rather than a
baseline file, as that way you can be sure that all your collection
parameters are exactly the same for the new Scanning Kinetics runs.
However, Good Laboratory Practice recommends that you collect a
new baseline for each laboratory session.
a Click Zero or choose Zero from the Commands menu.
b If you like, change the name of the blank/s.
c Insert blank samples into the appropriate positions of the
Multicell Holder and click OK.

How To…
15 Start the run

a Click the Start button to start a data collection. Alternatively
you can choose Start from the Commands menu. Do not add
your active reagent at this time.
NOTE At this point, the system will display the „Save File‟ dialog box if you have
selected „Storage On (Prompt at Start)‟ on the „Auto Store‟ page of the „Setup‟
dialog box. If so, enter the file name for this Scanning Kinetics run in the „File
name‟ field and click „Save‟.

change the names of the samples.
click OK. The ‘Sync Start’ dialog box will be displayed.
e At the end of the run, determine the actual Stop Time by
observing the last value in the ‘Time’ column of the ‘User
Data Form’.

How To…
Perform a Simple Reads measurement at a single wavelength

This procedure describes how to perform a Simple Reads
measurement at a single wavelength.
1 Set up instrument parameters (wavelength, Y mode)
a In Simple Reads, click the Setup button or choose Setup
from the Menu line to display the ‘Setup’ dialog box.
b Select Read at Wavelength and enter the required
wavelength.
c Under ‘Y Mode’, select the ordinate mode you require. The
ordinate mode determines the way in which the photometric
value is measured and displayed in your report.
d Click OK. The instrument will change to the new wavelength.
Make sure that the sample compartment is clear (or you have a blank
in position) and click Zero. (Wait for the ordinate reading to reach
‘0’.)
3 Read the sample
a Insert the sample into the sample compartment. Wait while
the instrument changes to the specified wavelength. The
current wavelength is displayed in the Abscissa status
display at the top right of the application window.
b Click the Read button to perform photometric measurements
on the sample at the specified wavelength. The result appears
in the Report area and includes the ordinate reading
obtained and the wavelength at which the reading was
measured.
4 Print the results
You can print the contents of the Report area by clicking the Print
button.

How To…
Thermal
The following Thermal procedures are described:
 Performing a run and determine Tm using Derivative calculations
 Performing a run and determine Tm using Hyperchromicity
calculations
Perform a run and determine Tm using derivative calculations

This procedure describes how to perform a Thermal run using a
single temperature rate and automatically calculate Tm from the first
derivative.
Setup dialog box
In Thermal, click the Setup button, or choose Setup from the Menu
a In the ‘Wavelength’ field, enter the wavelength that you
c In the ‘Ave. (averaging) Time’ field enter the required value.
NOTE For slow changes in temperature (that is, slow ramp rates), it is recommended
that a long Ave. Time (for example, 2 to 3 seconds) be set.
d Enter the ordinate values in the ‘Y min.’ and ‘Y max.’ fields.

a Specify the ‘Collect Temperatures’ parameters:
b In the ‘Start °C’ field, enter the temperature at which you
want to start the data collection.

How To…
NOTE Once you click „OK‟ to close the „Setup‟ dialog box, the Cary will drive the
temperature to the specified Start Temperature.
c In the ‘Return °C’ field, enter the temperature to which you

want the Temperature Controller to drive at the end of the
data collection.
d Use ‘Monitor’ to specify whether the temperature will be
measured at the block or the probe.
e Select Simple Collect to set a single rate of temperature
change for the entire Thermal run.
f Enter the ‘Data Interval’, ‘Ramp Rate’, ‘End Temperature’
and ‘Hold time’. Select the stages where you want to collect
data in the ‘Collect Data’ column.
4 Set up the lamp source operation and data display options
when performing collections overnight or unattended for
long periods of time.
c Under ‘Display Options’, select ‘Individual Data’ to display
Select ‘Overlay Data’ to superimpose the collected data of
Click the Accessories tab and make sure that no accessories are
selected.
6 Set up the analysis to reduce measurement interference
Select Smoothing and nominate the smoothing ‘Interval’ and ‘Filter
Size’ to control the number of measurement points to be averaged.

How To…
7 Set up a derivative method of analysis

a Select Derivative to calculate the temperature at which 50%
of the DNA strands have separated (Tm).
b Select Autocalculate to automatically perform a derivative
calculation at the end of each run.
NOTE You can manually perform the derivative calculation, once the run is complete.
c Define the range over which to calculate and plot the first
derivative of the selected scan.
d Enter the Interval at which data is sampled in order to
perform a derivative calculation. A derivative calculation
involves the collection of points from a plot. Selection of
these points is dependent upon the data interval chosen. The
data points are then used to generate a separate trace from
which a derivative calculation is performed. The selection of
the Derivative Interval is dependent on the rate of data
collection. If data is collected every 0.1 °C then a derivative
interval of 0.1 °C will give the most accurate result.
e Enter the Filter Size you require for the derivative
calculation. The derivative uses a Savitzky Golay technique
where a number of points surrounding an individual point
are averaged to produce a new, smoothed point. For example,
a five point derivation uses the two data points before and
after each data point.
NOTE The larger the filter size, the more data points that are included in the filter for
smoothing.
f Enter the temperature that you want to start the calculation

in the ‘Low Calculation Limit’ field.
g Enter the temperature where you want to end the calculation
in the ‘High Calculation Limit’ field.

How To…
8 Set up other recalculations and normalization

a Select Calculation and enter or select an equation in the field
to perform a calculation.
b Select Correction and enter the ‘Correction Temperature’ to
perform a Thermal Expansion Correction.
c Select Normalize and enter the temperature at which you
would like the data to be normalized.
to include graphics in the generated report.
d Select the ‘Autoconvert’ option you require. If you select

the current folder.
Select Storage Off.
View menu to display information about your current reaction.

How To…
12 Finish setup
Once you are satisfied with your method setup, click OK to save any
changes made and close the ‘Setup’ dialog box. The Cary will start to
drive the temperature to the start temperature.
a Click Zero or choose Zero from the Commands menu. A
‘Loading Guide’ will be displayed.
b Place a blank solution in the cell that is in the light beam in
sample compartment and click OK. The system will perform
an instrument zero on the blank solution.
14 Equilibrate the sample
Make sure that the Cary has reached the start temperature and has
been at this temperature for 2 to 3 minutes. If you entered a hold
time in the ‘Stage 1 Hold’ field (Cary page), the system will wait and
equilibrate at this temperature for the hold time before proceeding
with the ramp. It is not necessary to wait to click the ‘Start’ button.
15 Start the run
menu. The system will display a ‘Loading Guide’.
b Load your sample into the front cell holder.
c Enter the name of your sample then click OK. The Cary will
wait for the specified hold time and then perform the run.
16 After the run
At the end of the run, depending on what you have set in previous
steps, the ‘User Data Form’ may appear. If it does, fill in the
appropriate entries in the columns provided for the sample and click
OK. The calculation equation will be displayed in the report.
Also, since ‘Autocalculate’ (under ‘Derivative’) was selected on the
‘Analyze’ page of the ‘Setup’ dialog box, a derivative calculation will
be performed at the end of the data collection.
The sample under investigation, with corresponding Tm values, will
be displayed in the Report area as well as parameters used to
generate the selected data file, temperature profile, and the method
used to contain the Tm value.

How To…
A plot will be displayed in the Graphics area indicating the original

temperature data and a plot of the first derivative of the selected
scan. The point at which the derivative plot peaks (that is, maximum
or minimum gradient on the original plot) is reported as the Tm value
and the original plot is marked with an arrow to indicate this point.
To view a plot of the collected data, choose Graph from the View
menu.
17 Save your data
a From the File menu choose Save Data As.
b In the ‘Files of Type’ field, click Batch.
c Enter the file name for this run in the ‘File name’ field.
d Click Save.
Perform a run and determine Tm using hyperchromicity calculations

This procedure describes how to perform a Thermal run with
multiple temperature ramps and determine Tm using a
hyperchromicity calculation.
Setup dialog box
In Thermal, click the Setup button or choose Setup from the Menu
a In the ‘Wavelength’ field, enter the wavelength that you
c In the ‘Ave. (averaging) Time’ field, enter the required value.
NOTE For slow changes in temperature (that is, slow ramp rates), it is recommended
that a long Ave. Time (for example, 2 to 3 seconds) be set.

How To…
d Enter the ordinate values in the ‘Y min.’ and ‘Y max.’ fields.

3 Set up the collect temperature parameters
a In the ‘Start °C’ field, enter the temperature at which you
want to start the data collection.
NOTE Once you click „OK‟ to close the „Setup‟ dialog box, the Cary will drive the
temperature to the specified Start Temperature.
b In the ‘Return °C’ field, enter the temperature to which you

want the Temperature Controller to drive at the end of the
data collection.
c Use ‘Monitor’ to specify whether the temperature will be
measured at the block or at the probe.
d Select Advanced Collect.
e Enter the number of changes in temperature rate that you
require in the ‘Number of Stages’ field.
NOTE An increase or decrease in the temperature must be a separate stage.
f Enter the ‘Data Interval’, ‘Ramp Rate’, ‘End Temperature’

and ‘Hold time’. Select the stages where you want to collect
data in the ‘Collect Data’ column.
4 Set up the lamp source operation and the data display options
when performing Thermal data collections overnight or
c Under ‘Display Options’, select ‘Individual Data’ to display
Or select ‘Overlay Data’ to superimpose the collected data of

How To…

a Ensure that you have the Thermostattable 6 x 6 Multicell
Holder accessory installed before starting the run.
b Select Use Cell Changer to enable the accessory.
c Click Select Cells, and select the cells you require from the
e Clear Blank Correction.
6 Set up temperature display
Under ‘Temperature Display’, select Block and Probe 1 to turn on
monitoring of the Multicell Holder block and one temperature probe.
7 Set up the RBA accessory
a Under ‘RBA’, select Use RBA.
NOTE If you are using a Series I RBA, you must configure it using the Align application
before starting any experiments.
b Select either ‘Attenuation’ and enter the Abs value or select

‘Position’ and enter the angle position.
8 Set up the analysis to reduce measurement interference
Select Smoothing and nominate the smoothing ‘Interval’ and ‘Filter
Size’ to control the number of measurement points to be averaged.
9 Set up your method of analysis
a Select Hyperchromicity to calculate the temperature at
which 50% of the DNA strands have separated (Tm).
NOTE The hyperchromicity calculation is performed once the run is completed.

How To…
b Select Van't Hoff Calculation to calculate a Van't Hoff plot

(Ln(K(T)) vs 1000/T(deg K)) and activate the
‘Hyperchromicity’ items.
c Select ‘Self Complementary’ if the DNA molecule being
analyzed consists of identical strands. Otherwise, select ‘Non
Self Complementary’.
d Enter the concentration in the ‘Total Conc of Strands’ field.
e Enter the ‘Molecularity’ (that is, the number of strands) of
the hybrid. For example, enter ‘2' if the DNA molecule being
analyzed consists of two strands.
f Select Δ G to perform Δ G (free energy) and K (equilibrium
constant) calculations.
g In the ‘Number of Temperatures’ field, specify the number of
temperatures at which you would like to perform the Δ G and
K calculations. The number set here is reflected in the
‘Temperature’ table below this field.
h Enter the required temperature in each cell of the
‘Temperature’ table.
10 Set up other recalculations and normalizations
Setup dialog box | Calculations page
a Select Calculation and enter or select an equation to perform
a calculation.
b Select Correction and enter the ‘Correction Temperature’ to
perform a Thermal Expansion Correction.
c Select Normalize and enter the temperature at which you
would like the data to be normalized.
11 Set up your reporting and printing requirements
to include graphics in the generated report.

How To…
specified printer as well as the Report area..
d Select the ‘Autoconvert’ option you require. If you select

the current folder.
14 Finish setup
Once you are satisfied with your method setup, click OK to save any
changes made and close the ‘Setup’ dialog box.
a Click Zero or choose Zero from the Commands menu. The
b Place the blank solutions in the correct positions in the
holder then click OK to zero the specified cells.
16 Equilibrate the samples
Ensure that the start temperature has been reached and held at this
temperature for 2 to 3 minutes. If you entered a hold time in the
‘Stage 1 Hold’ field (on the ‘Cary’ page), the system will wait and
equilibrate at this temperature for the hold time before proceeding
with the ramp, and it is not necessary to wait to click the ‘Start’
button.

How To…
17 Start the run

Click the Start button or choose Start from the Commands menu to
start the Thermal run. The ‘Save As’ dialog box will be displayed.
18 Set up file name and trace names for the data
a Enter the appropriate name for your run and click Save. The
b Place the solutions in the correct cell positions in the holder.
c Enter the name of each sample in the corresponding field
then click OK.
The Cary will wait for the specified hold time and then perform the
Thermal run.
19 After the run
At the end of the run, depending on what you have set in previous
steps, the ‘User Data Form’ may appear. If it does, fill in the columns
for each sample trace and click OK. The calculation equation will be
displayed in the report.
20 Hyperchromicity calculations
The Cary WinUV software will now prompt you to use the Thermal
ruler to define various limits for the hyperchromicity calculations.
a A prompt will be displayed asking you to select the
Associated DNA limits using the Ruler. Click OK. The cursor
changes to ruler mode.
b Position the mouse pointer on the Thermal curve at one end
of the linear region of the Associated DNA part of the curve
(before transition). Click and drag along the curve and
release at the other end of the linear region.
The software will automatically calculate a linear least
squares line between the selected points and extrapolate to
both ends of the melting curve.
c An ‘Associated DNA Limits’ dialog box will be displayed,
displaying the upper and lower limits that you selected with
the Thermal Ruler. Click OK to accept the calculated line.

How To…
d A prompt will then appear asking you to select the

‘Deassociated DNA limits’ using the Ruler. Click OK on this
prompt. The cursor changes to ruler mode.
e Position the mouse pointer on the Thermal curve at one end
of the linear region of the Deassociated DNA part of the
curve (after the transition). Click and drag along the curve
and release at the other end of the linear region.
The software will automatically calculate a linear least
squares line between the selected points and extrapolate to
both ends of the melting curve.
f A ‘Deassociated DNA Limits’ dialog box will be displayed
displaying the upper and lower limits that you selected with
the ruler. Click OK to accept the calculated line.
NOTE The Associated and Deassociated DNA Limits dialog boxes allow you to re-select
your limits if you are not satisfied with the linear least squares lines drawn in the
graph box. To do this, click „Retry‟ and select another region. If you wish to exit
from the calculation mode, click „Cancel‟. The ruler trace is removed from the
graph.
The Linear Region Associated and Deassociated correction points are displayed
in the Report area.
NOTE If the two extrapolated lines cross at any point, a warning will be displayed. Click
„Cancel‟ and repeat steps (a) to (f) above and re-select the linear regions. Click
„Ignore‟ to proceed with the calculation using the regions you have selected.
Using the lines calculated in the previous steps, the software will
automatically create a second graph box and display the calculated
Alpha curve.
g A prompt will be displayed asking you to select the range on
the Alpha curve that will be used for the Tm calculation. Click
OK to make your selection.

How To…
h Position the mouse pointer on the alpha curve at one end of

the linear region. Click and drag along the curve and release
at the other end of the linear region.
i An ‘Alpha Tm Limits’ dialog box will be displayed displaying
the selected limits. Click ‘Retry’ to reselect the limits, if
required. Click OK to accept the selected ranges. A linear
least squares line is drawn between the selected points and
the value of temperature where  = 0.5 is calculated to give
the Tm value. (The Tm value is automatically reported in °C
and K.)
If you have chosen the Van't Hoff Calculation method to calculate
Δ G and K:
j Using the data calculated in Step (i) above, the system
automatically creates a third graph box displaying a Van't
Hoff plot. A prompt will be displayed asking you to select the
required range on the Van't Hoff plot to calculate Δ G and K.
Click OK.
k Position the mouse pointer on the Van't Hoff curve at one end
of the linear region. Click and drag along the curve and
release at the other end of the linear region.
l A ‘Van't Hoff Limits’ dialog box will also appear, displaying
the upper and lower limits that you selected with the ruler.
Click OK to accept the calculated line. Alternatively, click
‘Retry’ to reselect the limits using the ruler.
NOTE If there is an error in the Van't Hoff calculation, you will be prompted to reselect
the linear region.
A linear least squares line is calculated between these points and is

extrapolated to both ends of the Van't Hoff plot. The slope and
intercept of the calculated line gives the enthalpy and entropy of the
hybrid formation (Δ H° and Δ S° respectively).
The software will automatically send the results, using the calculated
alpha curve, to the Report area including:
 K(Tm)experimental

How To…
 K(Tm)from first principles

 K(T Tm)experimental/K(Tm)from first principles
21 Step 20 will be repeated for each sample trace measured.
Perform a UV Dissolution run (Cary 50)

This procedure describes how to perform a UV Dissolution run using
a Cary 50.
1 Configure the instrument
a In UV Dissolution, click the Configure button.
b Double-click Instruments Configuration to expand the
‘Instrument Configuration’ group. Enter the details for the
instrument and any accessories to be used in your
experiment.
NOTE If you have already performed a UV Dissolution run, the settings for these fields
are stored in the pre-saved method or data file.
c Click OK.
2 Set up method parameters
NOTE Click „Connect‟ if the system is not online.
a Click Setup to display the ‘Method Setup’ window and create

a new method.
b Expand the ‘Method’ group on the left by double-clicking
Method.
c Click Method Setup.
d Enter a method name in the ‘Method Name’ field.
e Choose ‘Single Tester’ or ‘Dual Tester’.
f Select Allow Media Change if a media change is required.

How To…
g Select Use Fraction Collector if a fraction collector is

connected to the system.
h Select Dual Sample if multiple volumes are to be pulled using
the fraction collector.
i Enter the name of the analyst in the ‘Setup By’ field. Verify
the time and date provided.
j Enter the name of the reviewer in the ‘Approved By’ field.
Verify the time and date provided.
3 Set up sampling points
a On the left of the window, click Sampling Points.
b Under ‘Time’, enter the desired time point.
c Under ‘Increment time’, enter the desired increment time.
d Using the up/down arrows, set the number of desired time
points.
e Click Add Time Point to display the configured point details.
NOTE To delete a row, highlight the un-needed time point and click „Delete Row‟.
To clear all sampling points and start over, click „Clear Table‟.
TIP By using „Increment time‟ (the time between the time points) and „Time Points‟
(the number of points that you wish to add), the number of time points entered
will be added and the time between each of the points will be the time entered in
the „Increment time‟ field. By clicking „Add cycles‟, the number of time points
will be displayed.
4 Set up product information

a On the left of the window, double-click Product.
b Click Product Information.
c Enter a product name in the ‘Product Name’ field.
d Describe the product being tested in the ‘Notes’ field.

How To…
e If necessary, under ‘Sample Label 1’, ‘Sample Label 2’, and

‘Sample Label 3’, enter lot number, batch ID, company
information, department, and so on. These headings are
displayed in the report.
5 Set up media
a On the left of the window, click Media.
b In the ‘Media’ field, enter the type of media being used for the
UV Dissolution test. To use previously entered media, click
the down arrow and select the medium.
c In the ‘Media Information’ field, enter any applicable
information regarding the dissolution media.
d Enter appropriate information in the ‘Description’ field. To
use previously entered descriptions, click the down arrow
and select the description.
e In the ‘Media Volume’ field, enter the appropriate volume of
media (in milliliters). To use previously entered volumes,
click the down arrow and select the volume.
6 Set up media change
NOTE The „Media Change Parameters‟ page is present when „Allow Media Change‟ is
selected on the „Method Setup‟ page.
a On the left of the window, click Media Change.

b In the ‘Media Two’ field, enter the second type of media being
used for the UV Dissolution test.
c Enter appropriate information in the ‘Media Information’
and ‘Description’ fields.
d In the ‘Volume Removed’ field, enter the volume removed (in
milliliters).
e In the ‘Volume Replaced’ field, enter the volume to be added
(in milliliters).
f Click the Media Change Time point down arrow to select the
time point for the media change.

How To…
g Select Automated Media Change if an automated system is

in place to change to the second medium.
7 Set up the dissolution tester spindle
a On the left of the window, double-click Tester.
b Click Spindle.
c Click the Apparatus Type down arrow to select an apparatus
type.
d In the ‘Initial Spin’ field, enter the initial spin rate (in
revolutions per minute).
e In the ‘Initial Spin Duration field’, enter the initial spin
duration (in seconds).
f In the ‘Spindle’ field, enter the spindle speed (in revolutions
per minute).
g In the ‘Spin Tolerance’ field, enter the speed tolerance (in
percent).
h In the ‘Infinity Spin’ field, enter the speed of the infinity spin
(in revolutions per minute).
i In the ‘Spin Duration’ field, enter the spin duration (in
minutes).
NOTE „Infinity spin‟ and „Spin duration‟ are unavailable for Media 1 if „Allow Media
Change‟ has been selected on the „Method Setup‟ page.
NOTE Configure „Spindle Media 2‟ if a media change is required.
8 Set up temperature
a On the left of the window, click Temperature.
b If AutoTemp is installed, select AutoTemp.
c In the ‘Probes’ field, enter the number of temperature probes
installed.
d In the ‘Bath Temperature’ field, enter the desired bath
temperature (in degrees Celsius)

How To…
e In the ‘Vessel Temperature’ field, enter the desired vessel

temperature (in degrees Celsius).
f In the ‘Temperature Tolerance’ field, enter the temperature
tolerance (± °C).
g In the ‘Stabilization Delay’ field, enter a time (in seconds).
NOTE This time allows the media to equilibrate before sending temperatures to the
computer.
h In the ‘Log Intervals’ field, enter the intervals (in minutes) to

report the bath temperature.
i For an extended test, in the ‘Evaporation Rate’ field, enter
the evaporation rate (in milliliters per minute).
9 Set up sampling parameters
a On the left of the window, click Sampling Parameters.
b In the ’Prime Volume’ field, enter the prime volume (in
milliliters). The prime volume should exceed the amount of
drawn media necessary to fill the whole system.
c In the ‘Pause Time’ field, enter the pause time (in seconds).
The pause time is the delay time between the end of the
prime cycle and the beginning of ultraviolet analysis.
d In the ‘Purge Volume’ field, enter the purge volume (in
milliliters). Enter a value that is at least twice the amount
necessary to fill the whole system with air. This ensures that
all stranded media is properly expelled.
e If a fraction collector is installed, configure it with the
‘Fraction Collector’ options.
f In the ‘Waste Drop Volume’ field, enter the waste drop
volume (in milliliters). The waste drop volume is the amount
of volume expelled before the sample is collected.
g In the ‘Active Channels’ field, enter the number of active
channels.
h In the ‘Sample Volume field’, enter the sample volume.

How To…
i If replacement media is installed, in the ‘Replacement

Volume’ field, enter the replacement volume (in milliliters).
j If a syringe pump is installed, configure it with the ‘Syringe
Pump’ options.
k In the ‘Plunger Speed’ field, enter the plunger speed (in steps
per second). Samples with a high viscosity may require a
slower plunger speed.
l In the ‘Aspiration Dwell’ field, enter the aspiration dwell
time (in seconds). The aspiration dwell time is how long the
plunger stops after pulling a sample. Samples with a high
viscosity may require a longer aspiration dwell time.
m In the ‘Filter Type’ field, enter the filter type.
n Select Use Filter Changer if a filter changer is installed.
o In the ‘Cycles Per Filter’ field, enter the cycles per filter. This
value controls the number of times each filter is used before
it is discarded.
NOTE The number of filters to be replaced at each time point is defined by the number
in the „Probes‟ box on the Method > Tester > Temperature page beneath
„AutoTemp‟. Ensure this number reflects the desired amount of vessel positions.
If „AutoTemp‟ is disabled, it may be necessary to temporarily enable it to adjust
the number accordingly.
10 Set up standards selection

a On the left of the window, double-click Standards.
b Select Capsule Blank to prevent blank correction on
standard solutions.
c On the left of the window, click Standards Selection.
d Select Read Blanks Online to read a blank sample at each
time point.
e Select Read Standards Online to read a standard solution at
each time point.
f Select Calibrate Using Online Standards to read the
standards and use the readings in the calculations.

How To…
NOTE Leave this option unselected if using a single offline standard value or a multi-
standard calibration file.
g Select Read Post Cycle Standard to read a standard after the

samples have been analyzed at each time point.
h Select Use Running Mean or Bridged Mean to use multiple
standard readings to calculate %Dissolved and mg Diss.
NOTE „Bridged Mean‟ is enabled only if „Read Post Cycle Standard‟ is selected. Bridged
mean uses the average of the two standards analyzed at each time point.
„Running Mean uses the average of all standards measured to this point.
i In the ‘Active Component’ field, enter the component name.

j In the ‘Label Content’ field, enter the amount (in milligrams)
of drug product.
11 Set up standards information
a On the left of the window, click Standards Information.
b Select Check Standard if a secondary standard is prepared.
NOTE Measurement of the Check Standard is not possible using UV Dissolution if

„Read Standards Online‟ is unselected on the „Standards Selection‟ page.
Alternatively, you may use Simple Reads, Advanced Reads, or Scan to measure a
Check Standard.
c In the ‘Std Weight’ field, enter the standard weight (in

milligrams).
d In the ‘Std Volume’ field, enter the standard volume (in
milliliters).
e If applicable, enter the standard dilution in the ‘Std Dilution’
field.
f In the ‘Std Medium’ field, enter the standard medium.

How To…
g In the ‘Standard Tolerance’ box, enter an appropriate

‘%Variance’ value.
NOTE This option is enabled only if measuring a check standard.
NOTE Configure „Standards Information Media 2‟ if a media change is required.
12 Set up standard calibration
NOTE „Standard Calibration‟ is unavailable if „Calibrate Using Online Standards‟ is

selected under „Standards Selection‟.
a Click Standard Calibration to use an offline standard value

or multi-standard calibration file.
b Select Single Standard if the absorbance is manually
entered. Enter the absorbance reading of the reference
standard for the active substance when it is 100% dissolved.
c Select Multi-standard if the linearity curve is required to
perform a UV Dissolution run. Click Browse to select a multi-
standard calibration file. The calibration file is stored with
the concentration method (*.mcn).
NOTE Configure „Standard Calibration Media 2‟ if a media change is required.
13 Set up control limits

a Click Control limits.
b Select Use Limits if standard control limits are required.
c Enter a value for the % difference in the ‘Difference’ field.
14 Set up spectrophotometer cell match
a On the left of the window, double-click Spectrophotometer.

How To…
b Click Cell Match.

c Select Use Cell Match Limits if cell match limits are
required.
d If ‘Use Cell Match Limits’ is selected, enter the absorbance
units in the ‘A.U.’ field.
15 Set up spectrophotometer pre-test
a On the left of the window, click Pre-Test.
b Select Do Pre-Test Scan to find the best wavelength for a
given standard.
c Enter an appropriate wavelength (in nanometers) in the
‘Scan From’ field.
d Enter an appropriate wavelength (in nanometers) in the
‘Scan To’ field.
e Enter the average time (in seconds) in the ‘Ave. (averaging)
Time’ field. Ave. Time corresponds to the number of flashes
for which the spectrophotometer averages the collected
signal. The longer the average time, the longer the signal is
averaged, resulting in one data point. Increasing the average
time results in a smoother scan. If the samples are highly
absorbing or turbid, use a longer average time to improve
accuracy by reducing the noise in the data.
f Enter an appropriate interval (in nanometers) in the
‘Interval’ field. Interval corresponds to how often scanning
occurs between the wavelength entered in the ‘Scan From’
and ‘Scan To’ fields.
16 Set up analysis (three options)
Single reading
a On the left of the window, click Analysis.
b Select Single Reading to perform an analysis at a fixed
wavelength.
‘Wavelength’ field.

How To…
d Enter an average time (in seconds) in the ‘Average Time’

field. Use Ave. Time to set the signal averaging time to be
used during data collection. If your samples are highly
absorbing or turbid, use a longer average time to improve
accuracy by reducing the noise in your data.
OR Scan
b Select Scan to perform a scan of the solution.
c Select Mean or Second derivative for an operation. If
‘Second Derivative’ is selected, enter an appropriate wavelength
(in nanometers) in the ‘Wavelength’ field.
e Enter an appropriate wavelength (in nanometers) in the
f Enter the average time (in seconds) in the ‘Ave. (averaging)
Time’ field.
g Enter an appropriate interval (in nanometers) in the
‘Interval’ field.
OR Single and scan
a Click Analysis.
b Select Single and Scan to perform an analysis at a fixed
wavelength and to scan the solution.
d Enter the averaging time (in seconds) in the ‘Ave. Time’ field.
f Enter an appropriate wavelength (in nanometers) in the
g Enter the averaging time (in seconds) in the ‘Ave. Time’ field.

How To…
h Enter an appropriate interval (in nanometers) in the

NOTE Configure „Analysis media 2‟ if a media change is required.
17 Set up correction (two options)

Single reading
a Click Correction.
b Select Do Correction to correct the reading at the analytical
wavelength.
c Select Single Reading.
d Enter the appropriate wavelength (in nanometers) in the
e Enter the averaging time (in seconds) in the ‘Ave. Time’ field.
OR Correction scan
a On the left of the window, click Correction.
wavelength.
c Select Scan.
f Enter the averaging time (in seconds) in the ‘Ave. Time’ field.
NOTE Configure „Correction Media 2‟ if wavelength correction and a media change is

required.

How To…
18 Set sample limits

a On the left of the window, click Sample Limits.
b Select Use Limits to incorporate limits for the sample time
points.
To remove any of the time points, clear the check box.
To reinstate all of the time points click All.
c Enter the Low (%Diss) and High (%Diss) for each time point.
d Click Default Table to reset the %Diss values.
19 Set report print options
a On the left of the window, double-click Reports, then click
Print Options.
b Select AutoPrint to print a report automatically after data
collection.
c Select User Data Form to enter extra information about the
traces in the graph box.
d Select Company Logo to include your company's logo in the
report.
e Select Parameters to include method setup parameters in
your report.
f Select System Configuration to include the settings and
configuration of your system (including your
spectrophotometer, pumps, vessels and testers) in your
report.
g Select Test Log to include a copy of the test log in your
report.
h Select the ‘Graph Type’ to include any of the graphs in the
respective report.
NOTE Adjust the X and Y scales to change the size of the graphs that appear on the
report. The default setting is X=40, Y=20. An alternate size may affect the
headings of certain graphs.

How To…
20 Set up report data

a On the left of the window, click Data.
b Select Absorbance to print the absorbance data for each
vessel for each time point specified in the Dissolution run. No
calculations are performed on this data. Enter the number of
decimal places to be displayed in the report in the ‘Decimal
Places’ field.
c Select ‘Uncorrected’, ‘Cell Match Corrected’, or ‘Cell Match
and Blank Corrected’ under ‘Absorbance Report Options’.
NOTE This option affects only the displayed data. All calculations are performed using
fully corrected values.
d Select mg Diss to print the calculated mg dissolved value for

each vessel at each time point specified. Enter the number of
Places’ field.
e Select %Dissolved to print the %Dissolved value achieved at
each specified time point for each vessel. Enter the number
of decimal places to be displayed in the report in the
‘Decimal Places’ field.
f Select Time to %Dissolved to print the time at which the
specified % dissolved value is achieved for each vessel during
the run. Enter the number of decimal places to be displayed
in the report in the ‘Decimal Places’ field.
g Select Temp/Stir Profile to include the bath temperature, in
degrees Celsius, and stir rate, in revolutions per minute, for
each time point specified. Enter the number of decimal
places to be displayed in the report in the ‘Decimal Places’
field.
h Select Statistics to print the calculated mean of all the
standard absorbance readings during the run, as well as
statistical data on these readings. This function includes Abs,
Time, %Diss and mg Diss statistics if these options are
requested for the report.

How To…
The statistics that are reported include: mean value (Mean),

the minimum (Min.) and maximum (Max.) standard
absorbance reading, as well as the Standard Deviation (SD)
and percentage Relative Standard Deviation (%RSD) for
every time point.
i If ‘Statistics’ is selected, you can enter the number of decimal
places to be displayed in the report for the SD and %RSD
options.
j Select the vessels you want to include in the report.
21 Set up report time points
a On the left of the window, click Time Points.
b Select All if all time points are to be reported.
c The ‘Time Points’ table displays by default the time points of
your data collection, corresponding to the collect timing
designated in ‘Sampling Time’ points. Select individual time
points by selecting the box next to them.
22 Set up report %Dissolution points
a On the left of the window, click Dissolution Points.
b Select All if all %dissolution points are to be reported. If not
all of the %dissolution points are required, clear the
unnecessary %dissolution points. The maximum number of %
Dissolved points you can enter for report purposes is 20.
23 Set up graph display
a On the left of the window, click Graph Display
b For each section, select Auto Scale Y Min and Auto Scale Y
Max to autoscale the graph(s). Otherwise, enter a value for
each field to define the graph axes.

How To…
Perform a UV Fiber Optic Dissolution run (Cary 50)

This procedure describes how to perform a UV Fiber Optic
Dissolution Run using a Cary 50.
1 Configure the instrument
a In UV Fiber Optic Dissolution, click Configure.
b On the left of the window, expand ‘Instruments
Configuration’ by double-clicking Instruments
Configuration.
c Enter details for the instrument and any accessories to be
used in your experiment. Click OK.
NOTE If you have already performed a UV Fiber Optic Dissolution run, the settings for
these fields will be stored in the pre-saved method or data file.
2 Set up method parameters
NOTE Click „Connect‟ if the system is not online.
a Click Setup to display the ‘Method Setup’ dialog box and

create a new method.
b On the left of the window, double-click Method.
c Click Method Setup.
d Enter a method name in the ‘Method Name’ field.
e Choose ‘Single Tester’ or ‘Dual Tester’.
f Select Allow Media Change if a media change is required.
g Enter the name of the analyst in the ‘Setup By’ field. Verify
the time and date provided.
h Enter the name of the reviewer in the ‘Approved By’ field.
Verify the time and date provided.
3 Set up sampling points
a On the left of the window, click Sampling Points.

How To…
b Enter the desired time point under ‘Time’.

c Enter the desired increment time under ‘Increment Time’.
d Using the up/down arrows, set the number of desired time
points.
e Click Add time point to display the configured point details.
NOTE To delete a row, use the up/down arrows under „Go To Row‟ to select the
appropriate row, and then click „Delete Row‟.
To clear all sampling points and start over, click „Clear Table‟.
TIP You can use „Increment time‟ (enter the time between the time points) and „Time
Points‟ (enter the number of points that you wish to add). Click „Add Cycles‟ and
the number of time points entered added and the time between each of the
points will be the time entered in the Increment field.
4 Set up product information

a On the left of the window, double-click Product.
b Click Product Information.
c Enter a product name in the ‘Product Name’ field.
d Describe the product being tested in the ‘Notes’ field.
e Under the ‘Sample Label 1’, ‘Sample Label 2’, and ‘Sample
Label 3’ fields, enter company information, department, and
so on, if necessary.
5 Set up media
a On the left of the window, click Media.
b In the ‘Media’ field, enter the type of medium being used for
the UV Fiber Optic Dissolution test. To view previously
entered media, click the drop-down arrow and select the
medium.
c In the ‘Media Information’ field, enter any applicable
information regarding the dissolution media.

How To…
d Enter the appropriate information in the ‘Description’ field.

To view previously entered descriptions, click the drop-down
arrow and select the description.
e In the ‘Media Volume’ field, enter the appropriate volume of
media in milliliters. To view previously entered volumes,
click the drop-down arrow and select the volume.
6 Set up media change
a On the left of the window, click Media Change.
NOTE The media change parameters page is enabled when „Allow Media Change‟ is
selected on the „Method Setup‟ page.
b Under the ‘Media Two’ field, enter the second type of media
being used for the dissolution test.
c Enter the appropriate information in the ‘Media Information’
and ‘Description’ fields.
d In the ‘Volume Removed’ field, enter the volume removed (in
milliliters).
e Under the ‘Volume Replaced’ field, enter the volume to be
added (in milliliters).
f Click the Media Change Time point drop-down arrow to
select the time point for the media change.
g Select Automated Media Change if an automated system is
in place to change to the second medium.
7 Set up the dissolution tester spindle
a On the left of the window, double-click Tester.
b Click Spindle.
c Click the Apparatus Type drop-down arrow to select an
apparatus type.
d Enter the initial spin rate (in revolutions per minute) in the
‘Initial Spin’ field.
e Enter the initial spin duration (in seconds) in the ‘Initial
Spin Duration’ field.

How To…
f Enter the spindle speed (in revolutions per minute) in the

‘Spindle’ field.
g Enter the speed tolerance (in percent) in the ‘Spin Tolerance’
field.
h Enter the speed of the infinity spin (in revolutions per
minute) in the ‘Infinity Spin’ field (unavailable if ‘Allow
Media Change’ is selected under ‘Method Setup’).
i Enter the spin duration (in minutes) in the ‘Spin Duration’
field.
NOTE Configure Spindle Media 2 if a media change is required.
8 Set up temperature
a On the left of the window, double-click Temperature.
b If AutoTemp is installed, select AutoTemp.
c Enter the number of probes installed in the ‘Probes’ field.
d Enter the bath temperature (in degrees Celsius) in the ‘Bath
Temperature’ field.
e Enter the vessel temperature (in degrees Celsius) in the
‘Vessel Temperature’ field.
f Enter the temperature tolerance in the ‘Temperature
Tolerance’ (± °C) field.
g For an extended test, enter the evaporation rate (in
milliliters per minute) in the ‘Evaporation Rate’ field.
h Enter the intervals (in minutes) in the ‘Log Intervals’ field to
report the bath temperature.
9 Set up sampling parameters
a On the left of the window, click Sampling Parameters.
b Enter the desired ‘Probe return height’ (steps) using the
up/down arrow keys.
c Select Resident Probes to keep the probes in the dissolution
media throughout the test.

How To…
NOTE The „Probe return height‟ allows the temperature and optic probes to be raised to
a pre-determined method-specific height during sampling for analysis by the
Fiber Optic Detector.
10 Set up standards selection

a On the left of the window, double-click Standards.
b Click Standards Selection.
c Select ‘Measure Pre-Test Standards’ to read the standard
samples before the test. Select ‘Measure Post-Test Standards’
to read the standard samples after the test.
d Under ‘Standard Tolerance’, enter an appropriate
‘%Variance’ or ‘Abs Difference’ value.
NOTE This option is enabled only if measuring pre- and post-test standards.
e Select Bridged Standards to use the average of the pre- and

post-test standards to calculate % and mg dissolved.
f Select Read Post Run Blanks if a blank measurement is
desired at the end of the test.
g Under ‘Blank Tolerance’, enter an appropriate ‘%Variance’ or
‘Abs Difference’ value.
NOTE This option is enabled only if measuring post-run blanks.
h Enter the component name in the ‘Active Component’ field.

i Enter the amount of drug product (in milligrams) in the
‘Label Content’ field.
11 Set up standards information
a On the left of the window, click Standards Information.
b Select Check Standard if a secondary standard is prepared.

How To…
c Enter the standard weight(s) (in milligrams) in the ‘Std

Weight’ field.
d Enter the standard volume(s) (in milliliters) in the ‘Std
Volume’ field.
e If applicable, enter the standard dilution(s) in the ‘Std
Dilution’ field.
f Under ‘Standard Tolerance’, enter an appropriate
‘%Variance’ value.
NOTE This option is enabled only if measuring check standard.
g Enter the standard media in the ‘Std Media’ field.
NOTE Configure Standards Information Media 2 if a media change is required.
12 Set up standard calibration
NOTE „Standard Calibration‟ is unavailable if „Measure Pre-Test Standards‟ is selected

under „Standards Selection‟.
a On the left of the window, click Standard Calibration to use

offline standards.
b Select Single Standard if the absorbance is manually
entered. For each vessel, enter the absorbance reading of the
reference standard for the active substance when it is 100%
dissolved.
c Select Multi-Standard if the linearity curve is required to
perform a dissolution run. Click Browse to select a multi-
standard calibration file. The calibration file is stored with
the concentration method (*.mcn) or as a batch file (*.bcn).

How To…
NOTE Configure „Standard Calibration Media 2‟ if a media change is required.
13 Set up spectrophotometer cell match

a On the left of the window, double-click Spectrophotometer.
b Click Cell Match.
c Select Use Cell Match Limits if cell match limits are
required.
d If ‘Use Cell Match Limits’ is selected, enter the absorbance
units in the ‘A.U.’ field.
NOTE Unlike flow cells, the fiber optic tips that define the pathlength are not machine-
made. Therefore, cell match is for information only when using fiber optic probes
and tips. For this reason, individual probes are baselined and standardized.
14 Set up spectrophotometer pre-test

a On the left of the window, click Pre-test.
b Select Do Pre-test Scan to find the best wavelength for a
given standard.
e Enter the averaging time (in seconds) in the Ave. Time field.
Ave. Time corresponds to the number of flashes for which the
spectrophotometer averages the collected signal. The longer
the averaging time, the longer the signal is averaged,
resulting in one data point. Increasing the averaging time
results in a smoother scan. If the samples are highly
absorbing or turbid, use a longer averaging time to improve
accuracy by reducing the noise in the data.

How To…
f Enter an appropriate interval (in nanometers) in the

‘Interval’ field. Interval corresponds to how often scanning
occurs between the wavelength entered in the ‘Scan From’
and ‘Scan To’ fields. For example, if the data was collected
between 200 and 300 nanometers and the interval is set to 1,
the data would be collected every 1 nanometer.
NOTE The upper limit of the interval is 5 nanometers.
15 Set up analysis (three options)

Single reading
b Select Single Reading to perform an analysis at a fixed
wavelength.
d Enter an averaging time (in seconds) in the ‘Ave. Time’ field.
Use Ave. Time to set the signal averaging time to be used
during data collection. If your samples are highly absorbing
or turbid, use a longer average time to improve accuracy by
reducing the noise in your data.
OR Scan
a Click Analysis.
b Select Scan to perform a scan of the solution.
c Select ‘Mean’ or ‘Second Derivative’ for an operation. If
‘Second Derivative’ is selected, enter an appropriate wavelength
(in nanometers) in the ‘Wavelength’ field.

How To…

OR Single and scan
a Click Analysis.
b Select Single and Scan to perform an analysis at a fixed
wavelength and to scan the solution.
d Enter the averaging time (in seconds) in the ‘Ave. Time’ field.
f Enter an appropriate wavelength (in nanometers) in the
g Enter the averaging time (in seconds) in the ‘Ave. Time’ field.
h Enter an appropriate interval (in nanometers) in the

required.
16 Set up correction (two options)

Single reading
a On the left of the window, click Correction.
wavelength.
c Select Single Reading.
d Enter the appropriate wavelength (in nanometers) in the
OR Scan
a Click Correction.

How To…

wavelength.
c Select Scan.
NOTE The mean of all absorbance values within the scan range will be used with the
Scan option.

required.
17 Sample limits
a On the left of the window, click Sample Limits.
b Select Use Limits to incorporate limits for the sample time
points.
c By default all time points are selected. Clear the check box
for each required time point.
d Enter the Low (%Diss) and High (%Diss) for each time point.
To clear the %Diss values, click Default Table.
18 Set up report print options
a On the left of the window, double-click Reports.
b Click Print Options.
c Select AutoPrint to print a report automatically after data
collection.

How To…
d Select User Data Form to enter extra information about the

traces in the graph box.
e Select Company Logo to include your company's logo in the
report.
f Select Parameters to include method setup parameters
within your report.
g Select System Configuration to include the settings and
configuration of your system (including your
spectrophotometer, pumps, vessels and testers) in your
report.
h Select Test Log to include a copy of the test log in your
report.
i Select the ‘Graph Type’ to include any of the respective
graphs in the report.
NOTE Adjust the X and Y scales to change the size of the graphs that appear on the
report. The default setting is X=40, Y=20. An alternate size may affect the
headings of certain graphs.
19 Set up report data

a On the left of the window, click Data.
b Select Absorbance to print the absorbance data for each
vessel for each time point specified in the dissolution run. No
calculations are performed on this data. Enter the number of
Places’ field.
c Select ‘Uncorrected’ or ‘Blank Corrected’ under ‘Absorbance
Report Options’.
NOTE This option affects only the displayed data. All calculations are performed using
fully corrected values.

How To…
d Select mg Diss to print the calculated mg dissolved value for

each vessel at each time point specified. Enter the number of
Places’ field.
e Select %Dissolved to print the % dissolved value achieved at
each specified time point for each vessel. Enter the number
of decimal places to be displayed in the report in the
‘Decimal Places’ field.
f Select Time to %Dissolved to print the time at which the
specified % dissolved value is achieved for each vessel during
the run. Enter the number of decimal places to be displayed
in the report in the ‘Decimal Places’ field.
g Select Temp/Stir Profile to include the bath temperature (in
degrees Celsius) and stir rate (in revolutions per minute), for
each specified time point. Enter the number of decimal
places to be displayed in the report in the ‘Decimal Places’
field.
h Select Statistics to print the calculated mean of all the
standard absorbance readings during the run, as well as
statistical data on these readings. This function includes Abs,
Time, %Diss and mg Diss statistics if these options are
requested for the report.
The statistics that are reported include: mean value (Mean),
the minimum (Min.) and maximum (Max.) standard
absorbance reading, as well as the Standard Deviation (SD)
and percentage Relative Standard Deviation (%RSD) for
every time point.
i If Statistics is selected, you can enter the number of decimal
places to be displayed in the report for the ‘SD’ and ‘%RSD’
options.
j Select the vessels you want to include in the report.
20 Set up report time points
a On the left of the window, click Time Points.
b Select All if all time points are to be reported.

How To…
c The ‘Time Points’ table displays by default the time points of

your data collection, corresponding to the collect timing
designated in Sampling Time points. Select individual time
points by selecting the box next to them.
21 Set up report %Dissolution points
a On the left of the window, click Dissolution Points.
b Select All if all %dissolution points are to be reported. If not
all of the %dissolution points are required, clear the
unnecessary %dissolution point ticks. The maximum number
of % Dissolved points you can enter for report purposes is 20.
22 Set up graph display
a On the left of the window, click Graph Display.
b For each section, select Auto Scale Y Min and Auto Scale Y
Max to autoscale the graph(s). Otherwise, enter a value for
each field to define the graph axes.
Set up tests for validation

The Validate application enables you to carry out a number of
performance tests to verify that the system is performing according
to specification. This procedure describes how to set up your
instrument to perform a validation test.
1 Set up test parameters
a In the Validate application, click the Tests menu or the Tests
button to display the ‘Configure’ page.
b Select the type of instrument performance tests that you
require. The various tests that are available will be displayed
as tabs across the top of this dialog box.
NOTE If your access privileges are restricted by the GLP application, you may find that
fields and buttons in this dialog box are not accessible.
c Click each tab and select the tests you require to be

performed.

How To…
d For each test, change any parameters that you require to be

changed to suit your laboratory.
NOTE If you wish to save the tests you have set up for future use, choose „Save
Method As‟ from the „File‟ menu to display the „Save As‟ dialog box and save the
method (with the file name extension „*.mvo‟).
2 Set up automated runs

If you are using either the Multicell Holder or the Auto Double
Aperture accessory to automate the testing procedure:
a Click Setup to display the ‘Setup’ dialog box.
b Click the Analyze tab.
c Select the appropriate accessory.
NOTE Check with your local Agilent office regarding the availability of this accessory.
3 Start the run

a Click the Start button or click Start on the Commands menu
to commence the testing procedure.
b If you set up for an automated run with the Multicell Holder
in the previous step, then the ‘Cell Loading Guide’ will be
displayed. Place the specified solutions and/or filters exactly
as shown in this dialog box and click OK.
Alternatively, follow the instructions on the prompts that
appear and click OK to continue with the performance tests.
Click Cancel at any time to stop the Validate run.

How To…
NOTE For manual runs (without the use of automated accessories), the Validate
application will automatically arrange the tests so that tests requiring your
intervention (for example, placing filters or solutions into the sample
compartment) are performed early in the Validate run. Tests that do not require
your intervention will be performed later in the run. The system will advise you
when you are no longer required. However, if you require, you can change the
order of the tests.
The results for each test are displayed in the Report area
immediately after the test is completed.
4 After the run
At the end of the Validate run, the Cary system automatically
generates a report file. The report file is stored in the Cary WinUV
directory.
The Report file will be saved with the following format:
DATE TIME.RVO
For example:
16 Apr 97 3;57;48 PM.RVO
The date and time will be displayed in the same form that is set in
the Windows Regional Settings Properties dialog box, accessed from
the Windows Control Panel.
NOTE At the end of the Validate run, you can also save your data. To do this, choose
„Save Data As‟ from the „File‟ menu to display the „Save As‟ dialog box and save
the current method, data and report in a batch file (BVO).

Troubleshooting
5. Troubleshooting
Instrument offline 154
Connect button instead of Start 154
Not enough memory 155
Poor display of videos and photographs 155
GLP log does not list some privilege changes 156
Report header and footer not being updated 156
To troubleshoot installation and startup problems, check the

information in this chapter. If you still have not found a solution to
your problem, check the ‘Troubleshooting’ section in the hardware
manual supplied with the instrument.
If you are having problems with your software, check the information
in this chapter to see if there is a solution to your problem. You may
also find a solution to your problem in the:
 ‘Troubleshooting’ section of the Help. To view this, click the
Windows ‘Start’ button, then ‘(All) Programs’, ‘Cary WinUV’, and
‘Cary Help’. Click ‘Troubleshooting’ and follow the links.
 Late Breaking News documents that were shipped with the Cary
WinUV software.
If you still have not found the solution to your problem, contact your
local Agilent office or representative.

Troubleshooting
Instrument offline
Problem
When I start the Cary WinUV software, the application reports that
the instrument is ‘Offline’.
Solutions
 Check the connection of the main instrument cable (IEEE cable)
attaching the computer to the instrument.
 Ensure the instrument has completed its initialization tests
before you start the Cary WinUV software.
 Contact your local Agilent office.
For more information, refer to Chapter 4 in the Hardware manual
provided with the instrument.
Connect button instead of Start

Problem
When I start the Cary WinUV application, I want to use the ‘Start’
button but it has changed to a ‘Connect’ button — why?
Solution
When you first start the Cary WinUV software, the System
Information application has control of the instrument while it
initializes. Until the initialization has finished, any other application
that you start will not have control of the instrument; hence the
‘Connect’ button. Wait until the ‘Connect’ button changes to ‘Start’. If
it does not change, check to see if you have any other Cary
applications running. If they are not collecting data, you can just
click the ‘Connect’ button to get control of the instrument.

Troubleshooting
Not enough memory

Problem
After installing version 3 of the Cary WinUV software, I get an error
saying that there is not enough memory and the Cary WinUV
software will not run.
Solution
Check that you have version 4.0 or later of Microsoft Internet
Explorer installed. (To check the version, start Internet Explorer and
choose ‘About Internet Explorer’ from the ‘Help’ menu. The version
number will be displayed).
Poor display of videos and photographs

Problem
The graphics and video display appears fuzzy, grainy or in weird
colors.
Solution
Increase your display settings to a higher resolution.
To do this:
1 Click the Windows Start button then (Settings), Control Panel
and double-click Display.
2 Click the Settings tab and adjust the color quality and screen
resolution to a higher setting (that is, 16 bit or 32 bit color),
providing your monitor and display adapter allow it.
3 Click OK.
NOTE You may have to restart your computer for the settings to take effect.

Troubleshooting
GLP log does not list some privilege changes

Problem
I have changed multiple privileges using the GLP application, but not
all of them appear in the log file that the application creates. What
has happened?
Solution
You need to click the ‘Apply changes now’ button on the ‘Privileges’
page each time you change privileges for a user or application. You
cannot make multiple changes and then click the ‘Apply Changes
now’ button — the changes are actually made but they don’t appear in
the log.
Report header and footer not being updated

Problem
I have changed the report header and footer information in the
System Information application, but it is not being updated when I
generate a report — why?
Solution
You need to be logged on as an Administrator to make the changes in
System Information.

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Cary WinUV

2 Cary WinUV Software Manual

Cary WinUV Software Manual 3

4 Cary WinUV Software Manual

Perform a temperature-controlled Enzyme Kinetics run

Cary WinUV Software Manual 5

Perform a run and determine Tm using derivative

6 Cary WinUV Software Manual

Cary WinUV Software Manual 7

Table 1. Applications included with each Cary WinUV software package

8 Cary WinUV Software Manual

Table 2. Descriptions of each Cary WinUV software application

Cary WinUV Software Manual 9

10 Cary WinUV Software Manual

Cary WinUV Software Manual 11

 Microsoft® or compatible mouse

12 Cary WinUV Software Manual

Preparation requirement Reference

NOTE If you are installing 21 CFR Part 11 software, refer to the:

Cary WinUV Software Manual 13

6 If prompted to complete installation of Cary WinUV by restarting

14 Cary WinUV Software Manual

8 Complete all the fields on the ‘Customer Details’ page. Click

Cary WinUV Software Manual 15

16 Cary WinUV Software Manual

NOTE If you are upgrading 21 CFR Part 11 software, refer to the:

 Cary WinUV Pharma Software Installation Instructions for 21 CFR Part

To upgrade the Cary WinUV software:

Cary WinUV Software Manual 17

10 Follow the prompts on the screen until the ‘Ready to Install’

11 If prompted to complete installation of Cary WinUV by restarting

13 Complete all the fields on the ‘Customer Details’ page. Click

18 Cary WinUV Software Manual

19 Insert the disk labeled Help.

Starting the software

Cary WinUV Software Manual 19

This page is intentionally left blank.

20 Cary WinUV Software Manual

Cary WinUV Software Manual 21

This chapter provides a brief introduction to the Cary WinUV

22 Cary WinUV Software Manual

Cary WinUV Software Manual 23

24 Cary WinUV Software Manual

NOTE GLP Administration is not present in UV Dissolution or UV Fiber Optic Dissolution

Cary WinUV Software Manual 25

Spectroscopy Configuration Manager

26 Cary WinUV Software Manual

The Privileges and Profiles software controls which

Cary WinUV Software Manual 27

Thermal (not for Cary 50)

UV Dissolution (Cary 50 only)

UV Fiber Optic Dissolution (Cary 50 only)

28 Cary WinUV Software Manual

Varian Spectroscopy Database Administrator

File name extensions

Cary WinUV Software Manual 29

The file name extensions for each file type are:

File type File name extension

where ** equals the Cary WinUV application code, used to distinguish