3 CCHM Trans Lec

CCHM 321: Clinical Chemistry 1│Lecture
2022-2023 3RD YEAR, 1ST SEMESTER Prof. Kimberly Ann Pulga, RMT, MPH
 2 Components of Quantitative Laboratory

Laboratory Chemicals, Result:
Supplies, and Equipment
(Preliminary Term, 3rd Topic) o Actual Value
Trans Outline: o Unit
Topic 1: Recap A. Glassware
A. Purpose of B. Special Glasses
 Système International d’Unités (SI)
Clinical C. Plasticware
Chemistry o Adopted internationally in 1960, is
D. Clinical
B. Role of Clinical Laboratory preferred in scientific literature and
Laboratory Supplies clinical laboratories and is the only
Scientist
C. Units of Measure
system used in many countries.
Topic 5: Laboratory
Topic 2: Chemicals and Vessels
o Based on the metric system
Reagents Topic 6: Pipettes
A. Analytical A. Types According
Chemicals to Calibration
B. Organic B. Blowout and
Reagents
Self-Draining
C. Reference
C. Types According
Materials
Topic 3: Water to Measure
Specification D. Types According
A. Processes of to Transfer
Purifying Water
Topic 7: Burets
B. Categories of
Water Topic 8: Syringes
Topic 9: Desiccator and
C. Types of Water
Desiccants
Purity
Topic 10: Balance
Topic 4: Laboratory Topic 11: Separation
Reagents and Supplies Techniques
RECAP
PURPOSE OF CLINICAL CHEMISTRY:
 To facilitate the correct performance of
analytic procedures that yield accurate and
precise information, aiding patient diagnosis
and treatment
ROLE OF CLINICAL LABORATORY
SCIENTIST:
 To correctly use basic supplies and
equipment and possess an understanding of
fundamental concepts critical to any analytic
procedure
UNITS OF MEASURE:
1
CCHM321 | BSMLS 2024 CLINICAL CHEMISTRY 1 TRANSCRIBER: CRUZ, A. M. A.
CHEMICALS AND REAGENTS

ANALYTICAL CHEMICALS:
 Analytic Reagent (AR) Grade
o Established by the American
Chemical Society (ACS)
o Labels on reagents state the actual
impurities for each chemical lot or list
the maximum allowable impurities
o Suitable for use in most analytical
laboratory procedure
o For qualitative and quantitative  United States Pharmacopeia (USP) and
analyses; essential for accuracy National Formulary (NF)
o Uses:
• Trace metal analysis o Used for manufacturing drugs that’s
• Preparation of standard why it is approved for human
consumption
o Limitations established for this group
of chemicals are based only on the
criterion of not being injurious to
individuals
o Pure enough for use in most chemical
procedures
o Purity standards are not based on the
needs of the laboratory
 Ultrapure (UP)
o Have been put through additional
purification steps
o Use in specific procedures such as
• chromatography
• atomic absorption
• immunoassays
• molecular diagnostics
• standardization
• other techniques that require
extremely pure chemicals
o Labels may carry HPLC – high
performance lipid chromatography
• Can be used for
chromatographic assays
2
 Chemically pure
o Approaches purity level of reagent
grade chemicals
 Spectroscopic
o Spectrally pure
 Chemically Pure (CP)  Chromatographic
o The impurity limitations are not o Has purity levels but they are attained
stated (difference with AR) by their respective procedures
o Preparation of these chemicals is not o Has minimum purity of 99%
uniform determined by gas chromatography
o Purity is usually delivered by
measurement of melting or boiling  Reagent grade
point o Contains impurity below certain
• Melting point analysis levels established by ACS
• Boiling point analysis
o Not recommended that clinical REFERENCE MATERIALS:
laboratories use these chemicals for  Primary Standard
reagent preparation unless there’s
o NIST - certified Standard Reference
• Further purification step
Materials (SRMs) / Certified
• Reagent black is included
Reference Material (CRMs)
• NIST – National Institute of
Standards and Technology
o Highly purified chemicals that can be
measured directly to produce a
substance of exact known
concentration and purity
o ACS purity tolerances for primary
standards are 100 ± 0.02%
 Secondary Standard
o Substance or chemical is of lower
purity with concentration determined
 Technical or Commercial Grade
by comparison with a primary
o Used primarily in manufacturing standard
o Should never be used in the clinical
laboratory WATER SPECIFICATIONS
ORGANIC REAGENTS:  Water is most frequently used reagent in lab

 Practical grade PROCESSESS OF PURIFYING WATER:
o Reagents with some impurities  Distillation
3
o Results in distilled water

o Almost all organic materials are
removed
o Water is boiled and vaporized
 Ion exchange/ Ion exchange resins
o Produces deionized water
o Percolates water through bead-like
spherical resin materials
o Ions in water are exchanged for other
ions that are fixed to the beads
 Reverse osmosis
o Pumps water across a semipermeable Ion exchange resins
membrane, produces RO water
• RO – reverse osmosis which
removes contaminants from
unfiltered water
o Does not remove dissolved gases
 Water can also be purified by ultrafiltration,
ultraviolet light, sterilization, or ozone
treatment
CATEGORIES OF WATER:
 According to CLSI, these categories of water
are based on specification needed for its use
rather than their method of purification and
preparation
A. Clinical Laboratory Reagent Water (CLRW)
Distillation
B. Special Reagent Water (SRW)
C. Instrument Feed Water
D. Water Supplied by Method Manufacturer
E. Autoclave and Wash Water
F. Commercially Bottled Purified Water
Reverse Osmosis
4
TYPES OF WATER PURITY: LABORATORY REAGENTS AND

 Type I SUPPLIES
o Used for test method requiring GLASSWARE:
minimum interference such as trace  High Thermal Resistant Glass
metal, iron, enzyme analyses
o Usually a low borosilicate glass with
o Consume immediately after it is low alkali content; borosilicate has:
produced
• has high degree of thermal
o Used for procedures that require resistance
maximum water purity for accuracy • most common type of
and precision glassware in volume
• Different procedures: measurement
preparation of standard
solutions, ultra o Resistant to heat, corrosion, or
microchemical analysis, thermal shock
flame photometry, atomic o Used when heating or sterilization is
absorption required
spectrophotometry, enzyme
o Brands: Pyrex and Kimax
analysis
o Most Common Resistant Borosilicate
 Type II
Glass
o Acceptable for most analytic
requirements, including reagent, • Beakers
quality control, and standard • Flasks
preparation
• Pipettes
o For qualitative chemistry procedures
and for most procedures done in
hematology, immunology,
microbiology and other clinical test
 Type III/Autoclave wash
o Glassware washing but not for
reagent preparation
o Usually used for urinalysis,
parasitology, and histology
 High Silica Glass

o Made by removing all elements from
borosilicate glass
5
o Has good optical qualities,  Low Actinic Glass

temperature capabilities and is
radiation resistant o Has materials that usually impart red
color to the glass that reduce the
o Used for high precision analytical amount of light passing through the
work and for optical reflectors and substance inside the glassware
mirrors
o Provides protection to reagents which
o Not used for the type of glassware are highly sensitive to light ranging
generally used in the laboratory from 3000 to 5000 Angstrom
o Glass made up of almost 96% silica o Used for substances that are
o Can be used at high temperatures particularly sensitive to light such as
(900-1200 degrees C) bilirubin and vitamin A
o Can stand a sharp change in
temperature
o Used for production of cuvettes and
thermometer as high silica glass is
considered optically pure
 Standard Flint Glass or Soda- Lime Glass

o Used to manufacture weighing bottles
because it develops less static surface
changes
 High Alkali Resistant Glass o Composed of a mixture of the oxides
o Partially used for strong alkali of silicon, calcium and sodium
solution o e.g. test tubes and weighing bottles
o Often referred as “soft glass” as its
thermal resistance is much less than
of borosilicate glass
6
SPECIAL GLASSES: o useful in making hot plates and table

 Colored and Opal Glasses tops
o Used in light fitters, lamp bulbs and  Radiation-Absorbing Glasses

lightning lenses
o Useful in preventing transmission of
huge energy radiation as gamma rays
and X-rays
PLASTICWARE:
 Begin to replace glassware in lab due to
notion that:
o It is much easier to use than glassware
o Inexpensive
o Disposable and prevent cross-
contamination
 Coated Glassware  Polyolefins
o Have thin metallic oxide permanently o Unique group of resins with relatively
fine- bonded to the surface of the inert properties (not reactive)
glass
o Unaffected by acids, alkali, salt
o Usually have electronic applications solutions and aqueous solution
as heat shield to protect against
infrared light o Can be autoclaved
o Two types
• Polypropylene – more
vulnerable to attack by
oxidizing agent; can
withstand higher temperature
• Polyethylene – Used
primarily to fabricate bottles,
beakers, jars, funnel, pipettes
connectors, and tubings; has 2
 Optical Glasses types:
o Mostly soda-lime, lead and • conventional
borosilicate of high optical purity • linear
o For making prisms, lenses, and
 Polycarbonate Resin
optical mirrors
o Twice as strong as polypropylene and
 Glass Ceramics (Pyroceram) maybe used at temperatures ranging
o Have high thermal resistance, 100 °C to 160 °C
chemical stability and corrosion
resistance like borosilicate glasses
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o Unsuitable for use with bases such as CLINICAL LABORATORY SUPPLIES:

amines, ammonia, alkaline and  Thermometers/Temperature – three major
oxidizing agents types of thermometers include:
o Dissolved by chlorinated aliphatic A. Liquid-in-glass thermometers
and aromatic hydrocarbons a. Use a colored liquid (red or other
o Used extensively in centrifuge tubes colored material) or mercury
and graduated cylinders b. Colored liquid is encased in
plastic or glass material
 Tygon c. With a bulb and a graduated stem
d. Three types
o Non-toxic, clear plastic of modified
i. Partial immersion
plasticized polyvinyl chloride
thermometers – usually
o Used to handle most chemicals but placed in heating blocks
should not be subjected to prolonged and water bath
immersion in aliphatic and aromatic ii. Total immersion
hydrocarbons, ketones and esters thermometers – for
refrigeration applications
o Flexible at 30 °C, brittle at 45 °C, and (storage)
resist dry heat to 95 °C iii. Surface thermometers –
o Can be steamed, autoclaved or may be needed to check
chemically sterilized temperatures for flat
surfaces such as incubator
o Used for manufacturing tubings and heating oven
 Teflon-Fluorocarbon Resin
o Pure translucent white and inert to
corrosive reagents, nitric acid, boiling B. Electronic thermometer or thermistor
chemicals, sulfuric acid, ketones, probe
esters and alcohols a. An accurate, fast-reading
o Can resist extreme temperatures electronic thermometer
ranging from -270°C to +255°C, used b. Advantages are size and
in cryogenic experiment or work at millisecond response time
temperature over extended periods c. Disadvantage: initial cost
o Used for self-lubricating stopcocks,

stirring bars, bottle cap liners and
tubing because of its anti-adhesive
properties
8
thermal stability should be

considered
b. designed to hold different
volumes rather than 1 exact
amount
C. Digital thermometer
LABORATORY VESSELS C. Graduated Cylinder

 Vessels holding or transferring liquid are a. Are long, cylindrical tubes
designed either usually held upright by an
octagonal or circular base
o To Contain (TC) b. Can be used to measure specified
volume of liquid
• Do not deliver that same c. Can be used when high degree of
volume when the liquid is
accuracy is not essential
transferred into a container
d. TD
o To Deliver (TD)
• Means that the labware will
deliver the same amount when
the liquid is transferred into a
container
A. Volumetric Flask
a. Is calibrated to hold one exact
volume of liquid (TC)
b. Has round, lower portion with a
flat bottom and long, thin neck
with an etched calibration line
PIPETTES
 Glass or plastic utensils used to transfer
liquids
B. Erlenmeyer Flasks and Griffin Beakers  usually used for volumes of 20 mL or less
a. Used in reagent preparation, flask  When using either pipet, the tip must be
size, chemical inertness, and immersed in the liquid to be transferred to a
level that will allow it to remain in solution
9
after the volume of liquid has entered the

pipet—without touching the vessel walls
 The pipet is held upright, not at an angle
 When transferring:
o Serologic/Mohr
• pipette should be held
vertically/at an upright angle
and the receiving vessel
should be steady
o Volumetric/Ostwald-Folin
• pipette is slightly tilted as well
as the receiving vessel
 Consider meniscus
o Stop at the bottom or lower meniscus
for clear/transparent solutions
o Use upper meniscus for colored
solutions
TYPES ACCORDING TO CALIBRATION:

 To contain (TC)
10
o Holds a particular volume but does o Allows the contents of the pipet to
not dispense that exact volume drain by gravity
o Calibrated by introducing the exact o Tip of the pipet should not be in
weight of mercury required to give contact with the accumulating fluid in
the desired volume at a specific the receiving vessel during drainage
temperature
TYPES ACCORDING TO MEASURE
 To deliver (TD)  Measuring pipets are used to transfer reagents
o Dispense the indicated volume and to make dilutions and can be used to
repeatedly to transfer a particular solution
o Calibrated by weighing the volume of
water that will flow from them by  Mohr Pipette
gravity o Does not have graduation to the tip
o Rate of delivery must never be o Self-draining pipet, but the tip should
hastened by blowing not be allowed to touch the vessel
 To blowout while the pipet is draining
o Same as TD pipette but drops o Calibration lies between two marks

remaining at the tip after delivery is on the stem
blown out of receiving vessel  Serologic Pipette
o An etched ring is seen near the mouth o Has graduation marks to the tip and is
piece generally a blowout pipet
• Has etched ring to indicate o Rate of fall is much too fast
that pipette needs to be blown
out o Has an etched band/ring on the
suction piece
 Between two marks
 Micropipette
o Calibration is affected by weighing
the water delivered between the two o Pipette with a total holding volume of
calibration marks less than 1 ml
BLOWOUT AND SELF-DRAINING o It may be designed as either a mohr or

PIPETTE: serologic pipette
 Blowout o To contain which is calibrated with
o Has a continuous etched ring or two mercury
small, close, continuous rings located TYPES ACCORDING TO TRANSFER
near the top of the pipette  Ostwald-Folin Pipette
o The last drop of liquid should be o Biologic fluids having a viscosity
expelled into the receiving vessel. greater than that of water
 Self-draining o They are blowout pipets, indicated by
o Without markings two etched continuous rings at the top
11
o Bulblike enlargement in the pipet • Air-displacement – relies on

stem a piston for suction creation to
draw the sample into a
o Measure smaller volume (2.0 mm or disposable tip that must be
less) changed after each use
o Dispense 1 volume without further
• Positive Displacement
subdivisions Pipette – operates by moving
 Volumetric Pipette the piston in the pipet tip or
barrel, much like a
o Designed to dispense or transfer hypodermic syringe
aqueous solutions and is always self-
draining • Dispensers and dilutor/
dispensers – automatic pipets
o Has the greatest degree of accuracy that obtain the liquid from a
and precision and should be used common reservoir and
when diluting standards, calibrators, dispense it repeatedly
or quality-control material
o Calibrated to deliver a fixed volume
of liquid
o Has a bulb between mouthpiece and
tip that decreases surface area and
diminish error from water film
 Pasteur Pipette
o Do not have calibration marks and are
used to transfer solutions or biologic
fluids without consideration of a
specific volume
Example of multi-channeled pipette used to
o Should not be used in any quantitative aspirate liquids with more than one volume
analytic techniques
 Automatic Pipette
o Advantages: include safety, stability,
ease of use, increased precision, the
ability to save time, and less cleaning
required
o May dispense a fixed volume or
variable volume
o Usually used in laboratory even in
clinical chemistry
o Three General Types of Automatic
Pipets
12
techniques such as chromatography or

electrophoresis
 Plunger is often made of a fine piece of wire
 Tips are not used when syringes are used for
injection of sample into a gas
chromatographic system
 Has parts including glass with fine barrels
 In electrophoresis work, however, disposable
Teflon tips may be used
Figure 1-7: found in automatic analyzers,
DESICCATOR AND DESICCANT
machine itself will pipette and dispense the
reagent  Hygroscopic
BURETS  Substances that take up water on exposure to
atmospheric conditions
 Wide, long, graduated pipet with a stopcock
at one end  Can remove moisture from the air as well as
from other materials – drying agents
 Total volume ranges from 25 mL to 100 mL
of solution o Desiccants are drying agents
 Used to dispense a particular volume of  Desiccants are most effective when placed in
liquid during a titration a closed, airtight chamber called a glass
desiccator
BALANCE
 Analytic and electronic balances are required
for the preparation of any primary standards
 The mechanical analytic balance is also
known as a substitution balance
 Has a single pan enclosed by sliding
transparent doors, which minimize
environmental influences on pan movement
 The weight range for certain analytic
SYRINGES balances is from 0.01 mg to 160 g
 Used for transfer of small volumes (usually
10 ml) in blood gas analysis or in separation
13
 A properly operating balance is essential in o Centrifuge classification is based on

producing high quality reagents and several criteria including
standards
• Benchtop or floor model
A. Electronic balances
a. Are single-pan balances that use • Refrigeration
an electromagnetic force to • Rotor head (e.g., fixed,
counterbalance the weighed hematocrit, swinging-bucket,
sample’s mass or angled)
b. Fast response time ( in less than
10 seconds, can already see the • Maximum speed attainable
mass of sample) o The speed of a centrifuge is easily
B. Analytical balance checked using a tachometer or strobe
a. Precision is up to 1/1000 grams light (once every 3 months)
C. Rough or Platform Balance
a. Precision is up to 0.1 gram o Used to obtain the sample
b. E.g. torsion balance and triple
o Should be switched off when not in
beam balance use
SEPARATION TECHNIQUES o Should be at full stop before opening
to prevent the sample from producing
 Centrifugation
aerosols
o Process in which centrifugal force is
o Note: will not be computing for RCF
used to separate solid matter from a
liquid suspension REFERENCES
o Centrifugation force depends on o Bishop, M. L., Fody, E. P., & Schoeff, L.
• Mass E. (2018). Clinical Chemistry: Principles,
Techniques, and Correlations. 8th ed.
• Speed Philadelphia: Wolters Kluwer
• Radius Health/Lippincott Williams & Wilkins.
o The speed is expressed in revolutions

per minute (rpm), and the centrifugal
force generated is expressed in terms
of relative centrifugal force (RCF) or
gravities (g)
o RCF 1.118 x 10-5 x r x (rpm)2
o Very common in clinical chemistry
 Centrifuge
o Consists of a head or rotor, carriers, or
shields that are attached to the vertical
shaft of a motor and enclosed in a
metal covering
14

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CCHM 321: Clinical Chemistry 1│Lecture

 2 Components of Quantitative Laboratory

CHEMICALS AND REAGENTS

 Chemically Pure (CP)  Chromatographic

ORGANIC REAGENTS:  Water is most frequently used reagent in lab

o Results in distilled water

TYPES OF WATER PURITY: LABORATORY REAGENTS AND

 High Silica Glass

o Has good optical qualities,  Low Actinic Glass

 Standard Flint Glass or Soda- Lime Glass

SPECIAL GLASSES: o useful in making hot plates and table

o Used in light fitters, lamp bulbs and  Radiation-Absorbing Glasses

o Unsuitable for use with bases such as CLINICAL LABORATORY SUPPLIES:

o Used for self-lubricating stopcocks,

thermal stability should be

LABORATORY VESSELS C. Graduated Cylinder

after the volume of liquid has entered the

TYPES ACCORDING TO CALIBRATION:

o Same as TD pipette but drops o Calibration lies between two marks

BLOWOUT AND SELF-DRAINING o It may be designed as either a mohr or

o Bulblike enlargement in the pipet • Air-displacement – relies on

techniques such as chromatography or

 A properly operating balance is essential in o Centrifuge classification is based on

o The speed is expressed in revolutions

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