ScanScope FL UserGuide PDF
ScanScope FL UserGuide PDF
ScanScope FL UserGuide PDF
Users Guide
Copyright Notice
Copyright 2007-2013 Leica Biosystems Imaging, Inc. All rights reserved.
Customer Resources
For the latest information on Leica Biosystems Aperio ePathology products and services, please visit www.LeicaBiosystems.com/ePathology.
Disclaimers
Use normal care in maintaining and using the eSlide Manager servers. Interrupting network connections or turning off the eSlide Manager and
DSR servers while they are processing data (such as when they are analyzing eSlides or generating an audit report) can result in data loss.
This manual is not a substitute for the detailed operator training provided by Leica Biosystems Imaging or for other advanced instruction. Leica
Biosystems Imaging Field Representatives should be contacted immediately for assistance in the event of any instrument malfunction. Installation
of hardware should only be performed by a certified Leica Biosystems Imaging Service Engineer.
ImageServer is intended for use with the SVS file format (the native format for eSlides created by scanning glass slides with the scanner).
Educators will use Aperio ePathology software to view and modify eSlides in Composite WebSlide (CWS) format.
Aperio ePathology products are protected by U.S. Patents: 6,711,283; 6,917,696; 7,035,478; 7,116,440; 7,257,268; 7,428,324; 7,457,446; 7,463,761;
7,502,519; 7,518,652; 7.602.524, 7,646,496; 7,738,688 and licensed under one or more of the following U.S. Patents: 6,101,265; 6,272,235;
6,522,774; 6,775,402; 6,396,941; 6,674,881; 6,226,392; 6,404,906; 6,674,884; and 6,466,690.
Customer Support
General Information
US/Canada/Worldwide Email:
TechServices@LeicaBiosystems.com
Email: ePathology@LeicaBiosystems.com
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Contents
1 Introduction to Scanning Slides...................................................................................... 6
2 Getting Started.................................................................................................................. 9
Prerequisites....................................................................................................................................9
Monitor Requirements....................................................................................................................9
Starting the Console........................................................................................................................10
What If the Scanner Is Not Connected to eSlide Manager?.................................................................... 11
Console Continue/Retry Window................................................................................................. 11
Selecting the Scanning Data Group..................................................................................................... 11
The Console Main Window................................................................................................................12
Viewing Slide Status........................................................................................................................15
Slide Operations Menu.....................................................................................................................17
Selecting Multiple Slides...............................................................................................................18
Slide Settings.............................................................................................................................19
Assigning a Parameter Set to a Group of Slides..................................................................................20
About Global Parameter Sets..........................................................................................................21
Error Handling................................................................................................................................22
3 Scanning Slides.............................................................................................................. 24
Connecting to the Aperio FL...............................................................................................................24
Loading Slides................................................................................................................................24
Automatic File Names...................................................................................................................27
Viewing Options..............................................................................................................................28
Setting up the Scan Area..................................................................................................................29
Channel Selection...........................................................................................................................31
Adding a Channel........................................................................................................................32
Channel Focus Offset Settings.....................................................................................................33
Changing Channel Scan Order.........................................................................................................33
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Contents
4 Snapshot Review............................................................................................................ 50
Getting Snapshots...........................................................................................................................50
Reviewing Snapshots.......................................................................................................................53
Locked Snapshots........................................................................................................................54
Scanning After Snapshots Are Reviewed..............................................................................................55
5 Configuration Options.................................................................................................... 57
Image Properties.............................................................................................................................57
Motion Properties............................................................................................................................58
Macro Move...............................................................................................................................58
Stage Move................................................................................................................................58
Stage Options.............................................................................................................................58
ScanScope Properties.......................................................................................................................59
Properties..................................................................................................................................59
Compression...........................................................................................................................62
Overwriting a Parameter Set Property...............................................................................................62
Parameter Sets...........................................................................................................................63
Parameter Sets Override Console Options......................................................................................63
Slide Settings.............................................................................................................................63
Area of Interest ..............................................................................................................................63
Defining Dyes.................................................................................................................................64
Defining a Filter Wheel Configuration...................................................................................................65
Defining a New Filter....................................................................................................................66
Removing or Moving a Filter...........................................................................................................67
Defining a Filter Cube Configuration.....................................................................................................67
Defining a New Filter Cube............................................................................................................69
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Contents
B Barcode Support............................................................................................................. 77
Barcode Support.............................................................................................................................77
Optical Character Recognition (OCR)....................................................................................................77
Prerequisites and Limitations..........................................................................................................77
Installing Barcode and OCR Support.....................................................................................................78
Barcode/OCR Options.......................................................................................................................78
eSlide Manager Barcode/OCR Support.................................................................................................80
Index...................................................................................................................................... 81
Symbols................................................................................................................................ 85
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Introduction to Scanning
Slides
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b. Position the blue diamond calibration point on a background area that does not appear to have tissue.
Calibration helps differentiate tissue from non-tissue and provides a flatfield correction for the camera. The
blue diamond calibration point must be located in the part of the slide that is under the coverslip. See Setting
up the Scan Area on page 29 for more on the calibration point.
c. Select the channels to tell the scanner what dyes (fluorochromes) are used for the sample and what filters to
use for those dyes.
d. Determine and apply the exposure times for each channel.
e. Place focus points (at least four) on various areas of the tissue. Focus points create a 3D map of your
specimen. Each focus point is assigned a value based on the perceived depth of the surface of the specimen at
that point. For example, a focus point on the peak of the tissue is a lesser number than a focus point in the
valley of the tissue. Based on the focus point values, the scanner determines the range of focus for the piece
of tissue, and adjusts the scanner objective while scanning to ensure the highest possible quality scan. See
Placing Focus Points on page 37 for more information on setting, moving, and adjusting focus points.
f. Perform calibration. Calibrating scans a small region near the blue diamond calibration point and uses this
image to obtain a shading correction image for the tissue. See Performing a Calibration on page 43 for
more information on calibrating.
9. Click the Start Scan button on the Console Scan tab.
When the scan is finished, the scan stripes for each channel are stitched together into a single, compressed image
and saved to your ImageServer or other location specified using the Console. You can now view the digital image of
the slide using the Console, ImageScope, or through eSlide Manager.
Note that pressing the red button on the front of the scanner halts the scan process and returns the slide stage to
the slide access position.
Images acquired on the Aperio FL using the quad multi-bandpass filter cube should be perfectly registered. Images acquired
using single-bandpass filter cubes or images imported from other sources may require registration changes so that the
channels are aligned correctly. For information on adjusting image registration, see the ImageScope Users Guide.
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See:
Magnification changer/doubler
Additional Manuals
For information on:
See:
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Getting Started
This chapter introduces the Console and contains information on basic concepts and features.
Prerequisites
The Aperio FL Console software is installed on the scanner control workstation by Leica Biosystems Imaging Technical
Services. If you wish to install the Console software on additional workstations, please contact Leica Biosystems Imaging
Technical Services for assistance.
Monitor Requirements
Because eSlides are by design high resolution and information rich, for best results you should use a high quality monitor to
view them. Make sure the monitor is at the proper viewing height and in a room with appropriate lighting. We recommend
any high quality LCD monitor meeting the following minimum requirements:
Display Type:
Screen Resolution:
Screen Size:
24-inch
Color Depth:
24-bit
Brightness:
300 cd/m2
Contrast Ratio:
500:1
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The first thing the Console does is ask you what scanner you want to connect to.
The Console passes along your login credentials if you open an eSlide from within the
Console by using another application (for example, by using the View Slide button to open the
eSlide in ImageScope), so you will not need to log in again to the new application.
2. Either type the name of the scanner you want to connect to in the Controller Name text box or select one from
the drop-down list of scanners that have been recently connected to.
3. Type your user name and password (the same user name and password you use to log into eSlide Manager). Your
eSlide Manager login must be assigned a user role that allows scanning.
4. Click OK.
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If you continue the scan, the scanned image is not cataloged by eSlide Manager, but it is saved on the scanner itself or, if
another location is defined as the local image share, in that location.
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The graphic on the Start tab differs depending on whether slides have already been scanned.
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The graphic may show that slides have already been scanned. An arrow indicates which slide in the tray is selected for
scanning.
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If slides are already loaded in the scanner and have already been scanned, the Scan tab may be selected instead of the
Start tab and the window may look something like this:
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Here are the color codes that can appear on the Start tab tray graphic:
Category
Snapshots
Slide State
Color Code
Light blue
Scanning
Green
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Category
Rack Contents
Slide State
Color Code
Orange
Red
White
The scanning status colors displayed on the Console Start tab graphic indicate how well the scanner was able to focus the
tissue on the slide. Although a well-focused slide is likely to result in a better image than a poorly-focused slide, status
colors should not be interpreted as an absolute indicator of image quality.
Instead, they should be used as a diagnostic tool alerting you to possible problems with a scan. Regardless of the scan
status color displayed by the Console, all slides should be reviewed for acceptable image quality.
For information on the colors that indicate scan quality and for tips on improving your scans, see Judging Scan Quality on
page 48.
You can also determine the status of a specific slide by holding your cursor over the slide in the Start tab graphic:
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`` Get Snapshot and Review Snapshot These commands relate to the snapshot review feature. See Chapter
4: Snapshot Review on page 50.
`` Save Slide Settings This command lets you quickly save a variety of settings (such as fluorochromes and
exposure times) that can be applied to multiple slides. See Slide Settings on page 19 for details.
`` Apply Slide Settings You can apply a saved slide setting to a group of slides. SeeSlide Settings on page
19 for details.
`` Assign Parameter Set You can apply a scanning parameter set to one or more slides. See Assigning a
Parameter Set to a Group of Slides on page 20.
`` Set Current Slide This command directs the scanner to start the next scan operation with this position and
continue scanning through all remaining tray positions then continue through the remaining tray positions in a
normal fashion. If a slide is currently being scanned, the scan will be completed before moving to the new current
slide position.
`` Rescan Slide Select this command to cause the scanner to rescan the slide after the current set of slides have
been completed.
`` Remove Slide Selecting this command will mark that tray position as empty. The Console automatically detects
empty positions and skips the empty slots while continuing scanning. You can save some time by manually marking
the empty slide positions so that the scanner does not go through the mechanical process of determining that they
are empty. You can also use this command if you want to leave the slide in the tray but dont want to scan it.
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`` Replace Slide Selecting this command will mark the slide position in the tray as occupied by a replacement
slide and ready for scanning during the next slide session.
`` Replace Rack Selecting this command will mark the entire tray as being replaced and ready for scanning during
the next slide session. If you lift out the tray, the tray status is reset just as if you used this command.
`` Clear Rack Marks every position in the tray as empty. Even if slides are loaded into the tray, they will be
skipped during scanning.
`` View Compressed Image If a compressed image is available, this command will open the scanned image file
in the ImageScope viewer.
`` View Uncompressed Image If an uncompressed image is available, this command will open the assembled
uncompressed image stripes in the ImageScope viewer.
You can also select specific slides using standard Windows techniques:
`` Click on one slide. Press and hold down the Shift key and click on another slide in the same tray to select a
contiguous group of slides.
`` Click on one slide. Press and hold down the Control key while you select non-contiguous slides.
`` Select multiple non-contiguous slides using the Control key, and then hold down the Shift and the Control keys to
select a contiguous group of slides without unselecting the previously selected slides.
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Slide Settings
Parameter sets (discussed in the next section) apply to overall scanner settings. Slide settings on the other hand, define
slide-specific settings such as channel configurations and exposure settings.
After saving slide settings, you can apply the same settings to multiple slides. For example, if you have a set of slides from
the same block in which the tissue area is the same, you can set the scan area on one slide, save the slide settings, and
then apply the settings to an entire group of slides.
To create a slide setting:
1. Right-click on a slide in the Start tab graphic and select Save Slide Settings. The Add Slide Settings window
displays:
2. Select the settings for that slide that you want to save. For example, if all slides use the same channel
configurations, select Channels and Exposure Settings.
3. Type a name for the slide setting in the Setting Name box.
4. Click Save.
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2. To remove a slide setting from a slide tray position, select that position, right-click, select Apply Slide Settings
and select None.
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In the example above, slides 3, 4, and 5 are selected (the black border indicates they are selected), so the parameter set
selected applies just to them.
Parameter sets are optimized to handle special situations. For an explanation of the different parameter sets available, see
Chapter 5: Configuration Options on page 57. Remember to set the parameter set to None when you are finished
scanning.
You can also use the Save Slide Settings and Apply Slide Settings commands to define and apply your own set of
parameters.
See Reviewing Snapshots on page 53 for information on setting a parameter set or slide setting for a single slide in the
Review Snapshots window.
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3. From Snapshot Review Selecting a parameter set here applies the parameter set only to the slide selected for
snapshot review and overrides global parameter settings.
Global ParmSet and None behave differently: selecting Global ParmSet returns the slide to using whatever global parameter
set has been applied, while None overrides any global parameter set, causing the global parameter set to be ignored for the
selected slides.
Note that one exception to this is the 2x3 parameter set; a snapshot review parameter set cannot override the fact that a
2x3 slide is being reviewed, but can override other settings such as compression.
Error Handling
Because a great deal of slide scanning is automated, if an error occurs during scanning, rather than halting the scanning
process the scanner logs the error and continues scanning. After the scan is complete, you can review the error log to see if
any errors occurred and then take action to repair any errors.
To view the error log, go to the View menu and select Error Messages. The Error Messages window appears:
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Scanning Slides
At any time you can bring up this window (for example, if your scanning session times out and you need to log in again or if
you want to switch to another scanner), by going to the Console ScanScope menu and selecting Connect.
Loading Slides
1. On the main Console window, click Manual Load.
The scanner slide tray is extended to provide access for slide loading.
2. Position slides into the tray with the coverslips on top and the slide labels oriented to the left. Ensure that the
slides are flat.
3. For the current slide, type the slide name in the File Name box and slide title in the Description box (both are
optional). (A large black arrow indicates which slide is the current slide. See the next page for an illustration.) If
you do not supply a file name, the scanner provides one.
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To automatically assign a file name based on the slides barcode, see Automatic File
Names on page 27.
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Tag
Replaced with
{ImageID}
{BarcodeID}
{Rack}
{Slide}
{Parmset}
You can also use the Console Configure window (see Chapter 5: Configuration Options on page 57) to set the file
name format permanently by using the ImageFilenameFormat property. The default value for that property is {ImageId}
simply replace it with your choice of the tags listed above, remembering to use the { } symbols. For example:
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Viewing Options
You can easily zoom the viewing window to the Area of Interest (AOI), full macro image, or the scan area by using the
controls at the upper left of the screen:
to expand it again.
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A simulated 10x scanning mode using 20x objective is also available. This mode uses a 20x objective at a faster
scanning rate to create a 10x scan.
If using the magnification changer (doubler), see Chapter 7: Working with a Magnification Changer on page
72.
2. Clear the Use Narrow Stripes check box to scan in wide stripes (1640 pixels wide) or select the Use Narrow
Stripes check box to scan in narrow stripes (992 pixels wide). Scanning wide stripes speeds up the scan. The
disadvantage of using wide stripes is that on uneven tissue, less tissue is focused. Narrow stripes provide better
focus with poorly prepared slides.
3. Select the tissue area to be scanned by adjusting the green selection rectangle using the following methods.
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a. Adjust the green selection rectangle area and location by clicking on the rectangle and dragging the
appropriate adjustment handles.
b. Alternatively, adjust the rectangle by entering area and position coordinates into the entry boxes located in the
Scan Area window. The Left field indicates the number of millimeters the selection rectangle is positioned
from the left edge of the slide viewing area. The Top field indicates the number of millimeters the selection
rectangle is positioned from the top edge of the slide viewing area. The Width and Height fields indicate the
width and height, respectively, of the selection rectangle (in millimeters).
The scan area must be within the Area of Interestyou cannot drag the scan area outside of the Area of
Interest.
4. Position the blue calibration point
symbol to an area that does not contain tissue (either inside or outside of the
scan area selection rectangle). It is crucial that the blue diamond not be located on tissue to achieve proper flatfield
(shading) correction.
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Channel Selection
Click the Channel Selection tab to view or define channels appropriate for the current glass slide.
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Adding a Channel
To add a channel:
1. Click Add Channel.
2. On the new channel line (shown in red on the Channel Selection tab), in the Dye column click Select a dye and
pick the dye to use from the list.
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6. If you prefer to enter the exposure time manually, you may do so by typing it in the Exposure column of the channel
table or using the arrows in the Exposure column. For details on automatically setting exposure, see Setting
Channel Exposure Time on page 34.
Channel Focus Offset Settings
Sometimes one channel has a slightly different focal plane than the first (reference) channel. This channel may require
a focus offset. To set a focus offset, type the value in the Focus Offset column. This value is relative to the registration
channel (the first channel listed on this window).
Multiple focus offset settings are available in the Console, and they are additive: Focus offset settings set by using the
global slide settings are added to the per-slide focus offset settings on the Focus tab (if any), which are added to the
channel focus offset settings on the Channel Selection tab (if any).
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1. Right-click on the macro image, select Adjust Exposure and then select an individual channel or All Channels.
2. If you have not already set exposure times, before finding the exposure time for the first channel, the scanner first
performs a macro focus and the Console then opens a window that shows the automatic exposure adjustment for
that channel at the spot you selected.
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3. The histogram at the bottom of the image shows the distribution of pixels for different intensity values. You can
see this distribution either on a linear or logarithmic scale by selecting the appropriate control on this window.
a. If you want to use this exposure, click Accept Exposure.
b. If you want to fine-tune the exposure, use the slider control on the left. As you move it up and down, the image
changes to reflect the change and the Reset Exposure and Check Exposure buttons are enabled. Note that
the updated image is an approximation of the new exposure. To verify it, you must use the Check Exposure
button.
c. To set the exposure back to the suggested value, click Reset Exposure.
d. If you have changed the exposure, you must click Check Exposure so that the Console can test this exposure.
When it is finished, click Accept Exposure if you are satisfied with the image.
The pixels shown in the histogram at the bottom of the exposure window should not extend into the red region. In
the example below, the red pixels in the image are saturated, indicating that this exposure time may result in an
image that is overexposed.
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The focus points you set appear as small yellow squares. If any of the focus points are positioned in clear areas of
the slide, you will need to delete them or reposition them away from the clear areas and into areas that contain
tissue. You can also add or remove focus points as needed. Larger scan areas should contain more focus points and
smaller scan areas should contain fewer focus points.
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`` To reposition a focus point, left-click it and then drag it to the desired location or select the focus point and use
the Move controls:
`` To remove a focus point, right-click it and then click Delete This Point from the context menu. To remove all
focus points, click Delete All Focus Points.
`` To add a focus point, double-click the new position or right-click the mouse in the desired location and select Add
Focus Point Here from the context menu. The context menu is slightly different if you right-click on a focus point
(as above) or right-click an empty position (below):
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1. Press the Alt and M keys or go to the View menu and select Video Monitor to open the Video Monitor window.
2. Resize the Video Monitor so it is wider than it is tall.
3. Drag the Video Monitor window so that it is adjacent to the macro image if needed.
4. Right-click one of the focus points and select Auto Focus This Point from the context menu.
Verify that the image in the Video Monitor window is in focus. The image you see is a direct feed from the line camera, a
series of stripes from the first channel. (All other channels are focused with respect to the first channel.) You want to see a
sharp image of the stripes. If the image is not sharp, use the focus sliders below the macro image to bring the image into
focus. Also, see Additional Focusing Tools on page 41 for more methods of focusing.
Note that the appearance of the stripes in the video window will vary depending on the tissue. If there are no stripes, then
your cursor is not on tissue or cells.
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If any focus points remain yellow, select them one at a time, reposition the focus point to a different area, rightclick, and then select Auto Focus This Point.
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The Focus Offset resets to zero when you load a new slide as this value is for a single slide only. For information on how
different focus offsets in the Console interact, see Channel Focus Offset Settings on page 33.
Moving the slider up moves the scanner objective up; moving the slider down moves the
objective down.
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`` You can also use the focus point context menu by right-clicking on a focus point:
`` Right-click on a focus point and select Manual Macro Focus from the context menu.
A Manual Macro Focus window appears on which you can use a coarse and fine sliders to fine-tune the focus of
the image in the video monitor window:
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`` Right-click on a focus point and select Clear All Focus Values from the context menu to reset all focus points to
their default value of 100. You will then need to use the Auto Focus button or manually focus the points before
performing a calibration.
Performing a Calibration
The Calibration operation acquires a shading correction image for each channel.
1. Click the Calibrate tab.
2. Click the Calibrate button.
A calibration image is acquired for each channel. When the calibration operation is complete, the calibration images
appears in the upper portion of the window. The calibration images should be relatively uniform from left to right with no
artifacts. (The illustration below shows three typical calibration images. The images will look different if you are using 20x
or 40x narrow stripes.)
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The calibration image for the first channel is displayed at the top with the other channels following in order. In the example
below, three channels have been defined.
The three calibration images may show different levels of darkness depending on the level of background fluorescence in
different channels, and this is not a problem. However, ideally each image should be even with no debris or artifacts, and
evenly shaded (not lighter or darker on one side than on the other).
If the calibration images display artifacts, go back to the Scan Area tab and move the blue diamond calibration point to
another location and make a new calibration image. Or you may want to use a default calibration image (see the next
section).
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If you want to use the default calibration image for this scan only, select the Use Default check box and click the Calibrate
button:
Calibration Reports
As part of their regulatory compliance process, some laboratories and clinics require a written record of equipment
calibration. The scanner calibrates itself each time you scan a slide and sends the calibration information to eSlide Manager
where you can access a complete report of calibrations. See the eSlide Manager Administrators Guide for details.
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Each channel is scanned separately in the same location (if you have three channels configured, then you see
the same stripe scanned three times, each time displayed in the color assigned to the fluorochrome used for that
channel).
The number of stripes in the scan is displayed during the scanning process. This number is based on the size of the
area to be scanned and whether the scan is in wide or narrow stripes.
3. After scanning is completed, click the View Slide button to preview the image (using ImageScope), to check the
quality of scan and focus. (See the next section for information on scan quality.)
At this point, the image is compressed in the SVS file format (or whatever file format or compression type you have
selected using the Console configuration optionssee Compression on page 62 for information on setting
different compression types).
All images are automatically saved and the scan information is logged into eSlide Manager after scanning is
complete.
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Successful Focus
Red
Failed Scan
Orange
Gray
Status is unknown
Yellow
In Progress
White
Empty
For example, the status display below indicates that the slide in position 2 of the slide tray was scanned with good focus.
We recommend reviewing all eSlides for quality, including those marked as Successful.
Additional status colors relate to the snapshot review feature. See Viewing Slide Status on page 3 for a complete list of
status colors.
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Snapshot Review
This chapter discusses using snapshot review to improve your slide-scanning workflow.
Operators can manually perform all of the pre-scan steps on one slide at a time.
In some cases, workflow is improved if the operator first views the snapshots of the slides, manually sets the scan areas,
sets focus points, chooses parameter sets, and so on, before scanning. The advantages of doing so are:
`` Smaller scan areas can be preselected by the operator, reducing the amount of time needed to scan the slides.
`` Snapshots of slides that are difficult to scan, such as slides containing high background staining, can be adjusted
before scanning, yielding better results.
`` Re-scans are reduced as the operator reviews the slide snapshots before the slides are scanned, spotting and
correcting potential problems before scanning.
`` If after review some scans are found to be of poor quality, snapshots of those slides can be reviewed and adjusted
to correct the problem before scanning those slides again.
For fluorescence scanning, snapshot review works best when all the slides use the same fluorochrome/filter combinations.
That way, you can create a single slide setting for the slides, apply that setting to all your slides, then use the snapshot
feature to set your scan areas, focus points, and so on.
Here is a summary of how snapshot review works (more details are below):
1. Load the slides you want to scan.
2. Choose a subset (or choose all) of the slides and tell the scanner to take snapshots of those slides.
3. Review the snapshots before scanning to set scan areas, focus points, scanning parameters, slide settings, and so
on.
4. After reviewing and adjusting all the snapshots, start scanning the slides.
Getting Snapshots
To get a snapshot, select one or more slides on the tray graphic on the Start tab of the console. (For information on how to
select more than one slide at a time, see Selecting Multiple Slides on page 18.)
After you have selected one or more slides, right-click with the cursor on one of the selected slides and select Get
Snapshot (or Get Snapshots if more than one slide is selected) from the context menu:
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While the slide is in the snapshot queue, the status color of the slide turns to white with blue diagonal lines. After the
snapshot is finished, the slide status color turns to light blue.
After it has been reviewed, the status color turns to dark blue. See the table below for all snapshot review status colors.
Here are all the slide status colors that apply to snapshot review:
Slide State
Color Code
Light blue
Dark blue
(See Viewing Slide Status on page 15 for information on all slide status colors.)
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Reviewing Snapshots
After snapshots have been made, select one or more slides on the Console Start tab graphic that display the light blue
status color and right-click while your cursor is over one of the selected slides. Select Review Snapshot (or Review
Snapshots if multiple slides are selected) from the context menu. The Review Snapshots window appears:
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`` Revert While you are making changes on the Review Snapshots window, the Revert button is enabled. Clicking
revert reverses all of the uncommitted changes you have made during this session.
If you make a change that changes a setting directly on the scanner, that change is considered committed and
cannot be reversed. For example, clicking Next commits all changes, so if you return to the slide by clicking Back,
the Revert button is disabled because the previous changes cannot be reversed.
The best way to understand this is to use the Review Snapshots window and to try the Revert button to see its
effect.
Locked Snapshots
If you try to review a snapshot that someone else is currently reviewing, the review screen has a yellow and black border
letting you know that the snapshot is lockedwhile you can look at the snapshot, you wont be able to change its settings:
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If you look at the Start tab, the yellow/black status color tells you which snapshots are being reviewed and are therefore
locked:
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To set another slide as the current slide, right-click on the new slide and select Set Current Slide from the context menu.
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Configuration Options
The Console Configure window allows you to view and change scanner settings.
To open the Configure window, go to the Tools menu and select Configure.
Image Properties
The Image Properties tab displays information about the current slide:
`` File Name The file name assigned to the slide by the person who scanned the slide.
`` Description The description may include details of the experiment or scanning conditions.
`` Update Changes the slide name and slide title to the values you type in the File Name and Description text
boxes. When rescanning a slide you may enter a new file name or description, but must click Update to commit the
changes.
`` Slide File Path The location of the slide image of the last scanned slide.
`` Scan Time The amount of time it took to scan the last slide and the area of the scan.
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Motion Properties
The Motion Properties tab contains options for moving the stage.
Macro Move
`` MC Move button Using macro camera coordinates, move the camera to the location specified by the x and y
values.
Stage Move
`` Stage Move Using stage coordinates, move the stage to the location specified by the x and y values.
`` Nudge by Moves the stage by the constant amount specified in the Nudge by box every time one of the four
nudge buttons is selected.
Stage Options
`` Extend Extends the stage to accept a slide.
`` Retract Retracts the stage.
`` Home Runs the stage homing procedure. Also puts the filter cubes and filter wheel in the home position.
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ScanScope Properties
The ScanScope Properties tab allows you to change options and view settings for the scanner.
Properties
1. To view a property setting, enter the Name and click Get.
2. To modify a property setting, enter the Name and a new Value and click Update.
Many of these properties are factory settings that should not be changed in the field. In fact, mixing these
settings improperly can have serious consequencesplease contact Leica Biosystems Imaging Technical
Services for advice if you are not sure how to use these properties.
Properties and parameter sets are stored in the controller.xml file, but we advise you to use the Console configuration
options to select/change them instead of editing the xml file directly.
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Here are some of the properties that you may wish to change or that Technical Support may advise you to change if they are
assisting you in troubleshooting a problem:
Property
Description
CacheSlideSettings
Saves the macro image of the slide in the tray. To enable, set to True.
CompressionQuality
Controls the JPEG and JPEG2000 compression quality. We recommend a value larger
than 20 and no more than 80. A quality factor of 30 typically results in acceptable image
quality at 30:1 compression (JPEG2000) or 15:1 compression (JPEG).
CompressionType
Settings: 0=None, 1=LZW, 2=SVS with JPEG, 5=SVS with JPEG2000 using Kakadu
library, 10=CWS, 12=JP2. See Compression on page 62 for more information.
DoCompression
Creates the file using the type of compression specified in CompressionType. For
example, if SVS is selected as the compression type, this parameter converts the file to
an .SVS file.
DoFastAlign
DoLabelCapture
Controls whether a label image should be captured during macro image processing.
DoWrite
DSRFailureResponse
Controls what happens if the scanner is not able to save the scanned image to eSlide
Manager because the connection to the DSR has failed. The two options are: 1) ask the
user if he wants to continue the scan saving the image locally or stop the scan (Ask)
or 2) continue the scan and save it locally without asking the user (Continue).
FiberLightIntensity
FileShare
The name of the Microsoft file share that points to the Stripes directory.
FocusOffset
GoodFocusThreshold
ImageLocation
The location where compressed slides are stored. Typically a network address.
ImageLocationByDate
If set to True, all compressed slides will be stored in ImageLocation in a folder with the
current date.
ImageServerPort
ImageServerURL
LCDPort
MacroAoiBottomPix
This number establishes the Area of Interest in which the search for tissue will occur.
This is the bottom coordinate where 0,0 is the upper left corner.
MacroAoiLeftPix
This number establishes the Area of Interest in which the search for tissue will occur.
This is the left coordinate where 0,0 is the upper left corner.
MacroAoiRightPix
This number establishes the Area of Interest in which the search for tissue will occur.
This is the right coordinate where 0,0 is the upper left corner.
MacroAoiTopPix
This number establishes the Area of Interest in which the search for tissue will occur.
This is the top coordinate where 0,0 is the upper left corner.
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Property
Description
MacroFocusPointRadius
Used by the tissue finder routines to control the number of focus points placed on
tissue.
MacroMaxNumberFocusPoints
Used by the tissue finder routines to control the number of focus points placed on
tissue.
MacroMinNumberFocusPoints
Used by the tissue finder routines to control the number of focus points placed on
tissue.
MacroNoiseErosionNumber
Used by the tissue finder routines to control the number of focus points placed on
tissue.
MacroSpatialSigmaThreshold
Used by the tissue finder routines to control the number of focus points placed on
tissue.
MacroUseBimodalNoiseReject
Used by the tissue finder routines to control the number of focus points placed on
tissue.
MacroUseColorOnly
Used by the tissue finder routines to control the number of focus points placed on
tissue.
MacroWhiteSigmaThreshold
Used by the tissue finder routines to control the number of focus points placed on
tissue.
PrescanDistance
PrescanDistance controls how far in millimeters the scanner scans when capturing a
calibration image. The default value is 1 millimeter. If you are having trouble getting a
good calibration image, increase to improve quality of the calibration image (but it takes
longer to get the image).
PrescanVelocityMultiplier
Decreasing this value can improve calibration images, but it takes longer to get the
image. Increasing this value can speed up calibration at the expense of image quality.
For slides with artifacts in the background (for example, slides prepared with tape
transfer), increasing this value can exaggerate those artifacts. If you are having trouble
getting a good calibration image, try setting this to 2 or 1 to improve the quality of the
calibration image.
QuickSnapAfterHome
If set to true, then the controller will take snapshots of all the slides in the tray after
starting up (homing). This can be useful when combined with the CacheSlideSettings
property. After the machine starts up, you see all the macro images in the Console. This
is only useful for scanners that use a multi-slide tray.
SaturationAdjust
Affects the Consoles video monitor. A value of 1 means do nothing; a value greater
than 1 makes colors richer; a value less than 1 makes colors duller.
SkipBlankStripes
Skips areas of the slide where there is no tissue. To enable set to True.
StripeLocation
TempImageLocation
If ImageLocation is unreachable, this location will be used temporarily. This must define
a valid disk path.
VSRPort
VSRServer
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Compression
By default, the scanner creates an eSlide using the SVS file format using compression type 2 (JPEG). You can use the
CompressionType to set the compression and file type for the image:
Setting
0
1
10
12
Compression
File Format
Extension
TIFF
TIFF
.TIF
.TIF
TIFF
.SVS
TIFF
.SVS
JFIF
.JPG
CWS directory
containing
multiple JFIF files
(directory)
JP2
.JP2
Please be aware that if you set compression type to 0 (uncompressed), you can potentially create an image file greater than
4GB, which Windows may not be able to open. If you have a need for uncompressed images, but they are too large for you
to use, contact Leica Biosystems Imaging Technical Services for alternative methods of creating uncompressed files that will
be easier to use.
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Parameter Sets
Parameter sets are predefined settings which have been provided for scanning certain types of tissue. By default, the
parameter set is None.
1. To change the parameter set, click on the arrow to the right of the Parameter Set box. You will see a list of
available parameter sets.
2. Select a parameter set from the list and wait for the Configure panel to update.
Here are some of the most commonly used parameter sets:
Parameter Set
Description
2 x 3 Support
This parameter set is covered in Chapter 6: 2x3 Slide Scanning on page 71.
None
When you have completed scanning with a parameter set, reset it back to None; leaving a parameter set active will cause
all future scans to be performed with the selected parameter set. Note that the Brightfield Alignment parameter sets are
used during the manufacturing process and should not be used in the field. You may define your own parameter sets
please contact Leica Biosystems Imaging Technical Services for assistance.
Parameter Sets Override Console Options
If a Console option setting is required for a particular parameter set, choosing that parameter will automatically override the
existing option settings.
Slide Settings
Choosing a slide setting from the Slide Settings drop-down box applies those settings to all slides in the slide tray. See
Slide Settings on page 19 for more.
Area of Interest
This window allows you to reposition the Area of Interest (AOI) by moving the red rectangle to suit your slide layout.
Note that the AOI does not indicate which area of the slide will be scanned, but rather which area will be considered for
scanning. The actual scan area must be a subregion within the AOI.
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In most cases you will get better results not using this feature, but instead performing a manual scan to let the Console set
the AOI.
Defining Dyes
On the Dyes tab you see the list of dyes currently defined for this scanner.
To search for a dye, begin typing the name of a dye in the Available Dyes search box. For example, to go to the first
occurrence of Alexa in the example above, type a in the search box.
You can also use the New Dye and Delete Dye buttons to define and delete dyes.
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2. Type the name of the dye you want to define and type the information about the dye (peak excitation and peak
emission values).
A color is automatically chosen based on that dyes peak emission wavelength. Select the display color for the dye
by entering an RGB value, using the slider to select a color, or clicking on a color box in the color display.
3. Click Save.
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2. Type the name of the filter (you might want to use its catalog number as its name), and band configuration
information.
3. Click Save.
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Multi-bandpass cubes will have a + symbol next to the position number. Click the + symbol to see all emission bands
included in the filter cube definition.
To see information on all emission bands included in this filter cube, click the filter cube on the map. For example:
Tip: To remove a filter cube, add a filter cube to an empty position, or change a filter cube in the scanner, you must move the
appropriate cube position in the turret to the loading position. To do this, select the filter cube position number in the Move
the filter number into load position drop-down list or double-click the position on the cube turret map shown on this tab.
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2. Type the name of the filter cube. You can use any name you like, such as dye name, catalog number, or description.
3. Define the emission and/or excitation filters included in this configuration. (For a multi-bandpass cube, click Add
Band to add a band.)
4. Click Save.
To add a filter to the wheel configuration after you have defined it, simply drag it onto the filter wheel map.
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The Aperio FL supports scanning slides that are 2x3 inches in addition to normal 1x3 slides. This option is an additional
feature and Aperio provides a separate slide tray which holds two 2x3 slides.
The settings required for 2x3 scanning are held as a Parameter Set within the Console.
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The ScanScope FL is provided with a built-in optical magnification changer (also known as a mag changer) that enables
scanning with different magnification using a single objective lens.
Magnification - for scanning with 20x Objective:
yyMagnification changer out = 20x
yyMagnification changer in = 40x
You will see on the ScanScope LCD system display that the mag changer is in. Pull gently until it stops before starting the
system.
To use the mag changer:
1. Push the mag changer in. If you are unable to push it all the way in, perform a manual scan and wait for the
macro image to appear; this will lower the objective lens.
The Console will automatically recognize the mag changer (doubler). The magnification will change and the bottom
left hand corner will show Scanner Ready, Doubler Inserted.
2. To return to the previous magnification, simply pull the mag changer out as far as it will go. The Console will return
to the original magnification.
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Troubleshooting
This appendix contains tips for troubleshooting the most common problems you may encounter with your
scanner.
If the information below does not help you find or fix the problem you are having, for further technical support contact Leica
Biosystems Imaging Technical Services. Refer to page ii of this manual for contact information. Also, see Troubleshooting
Tips on page 75 for more troubleshooting information.
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AppendixA: Troubleshooting
Thick Tissues
Thick tissues are difficult to scan. The Aperio FL is optimized for scans of samples less than 25. Although thicker samples
may be scanned, focus problems may occur.
Focus in thick tissues varies widely across the tissueit sometimes helps to scan using the Narrow Stripes setting.
Counterstains
DAPI is a popular counterstain that makes it easy to focus on the tissue because it stains the nuclei. DAPI may be included
in the mounting medium, but this may result in more background fluorescence. Therefore, some researchers prefer to stain
the slides with DAPI, then rinse off the excess dye before adding mounting medium and a cover slip.
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AppendixA: Troubleshooting
Troubleshooting Tips
Symptom
Solution
Verify that the power switch at the back of the unit is on and that the external
light is on.
If the stage is locked, you cannot scan. Contact Leica Biosystems Imaging
Technical Services for assistance.
1. Is the slide tissue unusually thick or thin? Try performing a manual macro
focus.
2. Is the label large or located inside the area of interest? Remove the label or
adjust the area of interest.
1. Try another slide to see if the problem is with the slide. Ensure that the
slide is not upside down in the slide tray. Move focus points to ensure that
tissue or cells are present for focusing.
2. Try performing a manual macro focus.
3. Unusually thick or thin slides may require adjustment of the macro focus
limits.
4. Another reason for not being able to obtain a macro focus is that the
exposure time for a channel is too short to give the scanner enough time
to find the macro focustry increasing the exposure time of the first
channel.
Verify that the slide is being held flat and not moving in the tray during scanning
(tape down the slide if necessary).
1. Assign additional focus points or move the focus points and repeat the
process.
2. Manually focus the focus points using the video monitor.
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AppendixA: Troubleshooting
Symptom
Calibration image or slide images are
not even or one side of each stripe is
less focused than the other side.
Solution
1. Calibration is probably of poor quality.
2. Ensure that there are no labels or debris on the underside of the slide. Slide
must be flat.
3. Calibration is critical to fluorescence scanningyou may need to use a
default calibration image. See Performing a Calibration on page 43 for
details.)
4. Repeat with another slide. An uneven calibration may also indicate optical
misalignment. This usually requires a service visit or factory repair.
1. Note whether exposure times for any of the channels is long (for example,
greater than .500 ms) and try adjusting them.
2. Use a default calibration image. (See Performing a Calibration on page
43 for details.)
3. Scan using narrow stripes.
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It may be that a motion fault occurred and was not recovered from. See the
Aperio FL Users Guide for information about recovering from a motion fault.
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Barcode Support
Glass slide labels often contain barcodes that contain information about the slide (for example, patient
name, case number, stain, etc.).
For licensed barcode types, the Aperio FL automatically reads the barcode on a glass slide label and saves the barcode
string with the eSlide. Human-readable text can also be read from the slide label using the Aperio optical character
recognition feature.
A one-time installation of a license enables support for all barcode types or OCR option for which you have purchased
support. (Customers who have previously purchased support for barcode types automatically receive support for the
previously purchased barcode types when they purchase a license for the new barcode types.)
Barcode Support
The Aperio FL supports the following barcodes: 2D DataMatrix, Interleaved (ID) 2 of 5, Code 39, Code 128, and QR Codes (a
two-dimensional matrix barcode that is common in Japan where it is the most popular type of two-dimensional barcode).
To purchase support for one or more barcode types, contact your Leica Biosystems Imaging representative, who will email
you a license string to decode the barcode types you have purchased support for.
Once the license string is installed, the scanner will automatically switch between licensed barcode types with no operator
intervention necessary when you scan slides that contain those types of barcodes.
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3. In the Name field, type BarcodeLicense and click Get to view the current setting.
4. In the Value field, type the license string you received.
5. Click Update.
Barcode/OCR Options
To provide support for marginal barcodes that may not meet specifications, the Aperio FL Console provides several
properties to help with the decoding of those barcodes. If you find you are using these properties frequently for your
particular barcodes, you may want to save them as a parameter set to make them easy to call up for your scanning.
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Property
Description
Values
Barcode Types
Supported
BarcodeUseShortMargin
True or False
BarcodePartialMinLength
BarcodeImproveBounds
True or False
BarcodeChecksum
ID 2 of 5, Code 39
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Property
Description
Values
Barcode Types
Supported
BarcodeC39FullASCII
Code 39
BarcodeDMSymbolSize
DataMatrix
True or False
ID 2 of 5, DataMatrix, Code
39, Code 128
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Index
A
Aperio FL Console. See Console
Area of Interest, AOI 25, 63
automatic file names 27
B
barcode support 77
C
calibration 7, 43, 44
default image 44
report 45
calibration point 25
channel exposure time 34
channel selection 31
compression 7, 62
configuration
image properties 57
motion properties 58
options 57
ScanScope properties 59
connect to scanner 10, 24
Console 8
Area of Interest (AOI) 28
connect to the scanner 10
global parameter 21
group parameters 20
installing 10
logging in 10, 24
macro image 28
main window 12
manual scanning 24
none parameter 21
scan area 28
scanning 55
slide status colors 15, 48
snapshots
getting 50
locked 54
reviewing 50, 53
starting 10
Start tab 12
status graphic 12
continue/retry when DSR connection lost 11
controller properties 60
D
data group, selecting 11
default calibration image 44
digital slide
file name 24
from barcode 27
doubler 72
dye
defining 64
setting display color 65
E
error
handling 22
log 22
eSlide
compression 7
creating 7
data group 11
eSlide Manager 7
exposure time, channel 34
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Index
F
filter cube
defining 67
deleting 70
moving 70
filter wheel
defining 65
deleting a filter 67
moving a filter 67
fluorochromes
dye 32
focus offset
additive offsets 33
per channel 33
per slide 41
focus points
adding, repositioning, moving 38
automatically focusing 41
manually focusing 42
placing 37
focus slider 41
assigning to snapshots 53
global 21
overwriting property 62
problems 73
R
regulatory compliance, calibration reports 45
narrow stripes 29
J
judging scan quality 48
L
live video monitor 39
loading slides 24
logging in 10, 24
M
magnification changer, using 72
manuals, list of 8
OCR support 77
P
parameter sets
assigning to multiple slides 20
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troubleshooting 73
U
uncompressed files 62
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Index
V
viewing scanned image 47
W
wide stripes 29
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Index
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Symbols
The following symbols may appear on your product label or in this users guide:
Manufacturer
Date of manufacture (year - month - day)
European Union Authorized Representative
In vitro diagnostic device
Serial number
Relative humidity range
Storage temperature range
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www.LeicaBiosystems.com/ePathology