Lecture (week2) Flashcards
preferred in
scientific literature and clinical laboratories and is the only system used in many countries
o Accepted internationally
o Provides global scientific community a uniform method of describing physical quantities
Système International d’Unitès (SI)
o Derivative or a mathematical function of one of the
basic units
o Example: m/s; still from base unit; can be
expressed using different parameter
Derived unit
o Widely used that they have become acceptable for
use with SI basic or SI derived units
Some Non-SI units
o When added to a given unit, it can indicate decimal
fractions or multiples of that unit
o Important note:
▪ The SI term for mass is kilogram or Kg. It is
the only basic unit that contains a prefix as
part of its naming convention
SI uses standard prefixes
- Electronic transmission of laboratory data and the more routine use of an electronic medical record, coding,
billing, and other data management systems may vary.
o Varies from one institution to another. Depends on
the SOP.
ELECTRONIC REPORTING OF RESULTS
- A substance that is used to test for the presence of another substance by causing reaction with it.
- Any substance employed to produce a chemical reaction
REAGENTS
substances that occurs naturally or is obtained through a chemical process. These analytical chemicals used in
the laboratory exists in varying grades of purity.
CHEMICALS
I. Analytical Reagent (AR)
II. Ultrapure
III. Chemically Pure (CP)
IV. United States Pharmacopeia (USP)
V. National Formulary (NF)
VI. Technical or Commercial Grade
CHEMICAL GRADES OF PURITY (INORGANIC REAGENTS)
o Highest grade of purity
o Established by ACS- American Chemical Society
o used for qualitative and quantitative analysis in
the laboratory
o used for special testing in the lab: trace metal
analysis and preparation of standard solutions
Analytical Reagent (AR)
o Additional purification which could be more sensitive
o Used for chromatography, atomic absorption spectrophotometry, molecular assays, molecular diagnostic for standardization, or other techniques
that require extremely pure chemicals
Ultrapure
o Pure grade
o Purity of the chemical is specified by the
manufacturers
o Chemically pure substances fail to reveal the tolerance limit of its impurities
o Not intended for research and analytical chemistry
Chemically Pure (CP)
o Pure enough to use in most chemical procedures however, it should be recognized that the purity standards are not based on the needs of the laboratory.
o May or may not meet all assay requirements.
o Good for human consumption
o Used in drug manufacturing
United States Pharmacopeia (USP)
o Same characteristics as USP
National Formulary (NF)
o Not used in clinical laboratory testing and only used in manufacturing industry
Technical or Commercial Grade
Also have varying grades of purity that differs from those used to classify inorganic reagents
Organic Reagents
o Practical grade with some impurities
o Chemically pure
o Spectroscopic
o Chromatographic
It approaches the purity level of reagent
grade
Chemically pure
Spectrally pure
Spectroscopic
The minimum purity of 99% could be
obtained only by gas chromatography
Chromatographic
o highly purified chemical that can be measured directly to produce a substance of exact known concentration and purity; purest
o ACS purity tolerance test for primary standard should be 100% ± 0.02%; equal to perfection
o Most biologic constituents are unavailable within
this limitation
Primary standard
Used in place of an ACS primary standard in clinical work and is often used to verify
calibration or accuracy of assessment/testing
NIST: Standard/Certified Reference Material (SRM/CRM)
o substance of lower purity with concentration determined by comparison with a primary standard
o prepared solution whose concentration is determined by testing
Secondary Standard
the most frequently used reagent in the
laboratory.
Water
o Distillation (distilled water)
o Ion exchange (deionized water)
o Reverse Osmosis
o Other purification processes: Ultrafiltration,
Ultraviolet Light, Sterilization, Ozone Treatment
Purification
o Classification according to CLSI as: Clinical laboratory reagent water (CLRW), Special reagent water (SRW), Instrument feed water, Water supplied by method manufacturer, Autoclave and wash water, and commercially
bottled purified water.
Reagent Grade Water
remove particulate matter for municipal
water supplies before any additional treatments
o Glass, cotton (to filter more particulate matter), activated charcoal (remove organic materials, even chlorine), submicron filters (≤ 0.2 mm; remove substances like bacteria)
Prefiltration
water has been purified to remove almost
all organic materials
o Water is boiled and vaporized.
o Water can be distilled more than once with each distillation cycle removing additional impurity
Distillation
water has some or all ions removed
o Although ions are removed, organic materials may
still be present
o If water goes ion exchange treatment, it is neither
pure nor sterile
Ion exchange
process that uses pressure to force
water through a semipermeable membrane
o Water that reflects a filtered product of the
original water is an RO water
o In RO, it does not remove dissolved gasses.
o At most cases, reverse osmosis may be used
as a pre-treatment of water.
Reverse Osmosis
excellent in removing particulate matter, microorganisms, and any pyrogens or
endotoxins
o Through ultraviolet oxidation, organic materials are removed. Sterilization processes, together with ozone treatment, removes bacteria, other microorganisms, or other residual products
Ultrafiltration & Nanofiltration:
Most pure
- ideal for special testing in the laboratory; trace metal determination, ion determination, enzyme analysis, tissue or cell culture, and the likes. Silicate content and resistivity goes hand in hand. The less
silicate content, the more the water is resistant. The more the water is resistant (less conductive), the more the water is pure.
Type I
still acceptable in most analytical requirements. Used in
quality control, reagent preparation, and laboratory determination.
Type II
Type III
not ideal for analysis or reagent preparation
Analytical reactions occur in optimal temperatures.
o Liquid in glass: usually measure temperatures between 20°C and 400°C
o Electronic / Thermistor probe: smaller and has millisecond response time
o Digital
Thermometers
are calibrated in the laboratory to make sure that accurate temperature is obtained. Calibrator: gallium
Thermometers
Differ on their ability to resist acid and alkaline solution and thermal resistance.
Glassware
▪ Ideal for heating and sterilization processes
▪ Has high thermal resistance
▪ Example: Pyrex, Kimax
Borosilicate Glass
▪ Soda lime glass
▪ Inexpensive but has low resistance to
temperature and chemical
▪ Easy to melt
▪ Usually, a disposable glassware (petri dish)
Flint Glass
▪ Silica glass
▪ Quite expensive
▪ Used when excellent light transmission is
needed
▪ Used for cuvettes (holds solution in
spectrophotometer)
Quartz Glass
▪ Soft glass
▪ High resistance to alkaline solution
Boron-free
▪ Corex
▪ Strengthened chemically than thermally
▪ Chemically-wise, 6x stronger than
borosilicate glasses
Aluminosilicate
▪ Used for high thermal requirement
▪ Could be heated up to 900⁰C
▪ Acid and alkaline resistant
Vycor
▪ Amber glasses; light protection
Low-actinic
Somehow begins to replace glassware in laboratory setting. Plasticwares are unique and high resistance supplies- resistance to corrosion and breakage.
Inexpensive compared to glassware.
Plasticwares
▪ Unique group of resins unaffected by acids,
alkaline, salt, and aqueous solutions
▪ Can be autoclaved
▪ Two types
* Polypropylene
* polyethylene
Polyolefins
▪ 2x stronger as compared to propylene
▪ Can resist temperature up to 100-160⁰C
▪ Dissolved in chlorinated aliphatic or aromatic
hydrocarbons
Polycarbonate resins
▪ Non-toxic, clear, can handle different
chemicals
▪ Can be steamed, autoclaved, or chemically
sterilized
▪ Flexible at around 30⁰C or brittle at around
45⁰C
Tygon
▪ Can resist extreme temperature of up
to -270⁰C to 255⁰C
Teflon-fluorocarbon resin
- Generally:
- Blood clots
- New pipets
- Metal ion determination
- Grease
- Bacteriologic
- Permanganate stains
CLEANING OF LABWARE
o Soapy water/ dilute bleach solution
o Acid dichromate
o 20% Nitric Acid
Generally:
o 10% sodium hydroxide
Blood clots:
o 5% hydrochloric acid
o 5% nitric acid
New pipets:
o 20% nitric acid
Metal ion determination:
o Potassium hydroxide and 10% ethanol
o Contrad 70
Grease:
o 2-4% cresol
Bacteriologic:
o 50% hydrochloric acid
o 25% sulfuric acid
Permanganate stains
o calibrated to hold one exact volume of liquid (TC)
o has a round, lower portion with a flat bottom and a
long, thin neck with an etched calibration line.
o Used to bring a given reagent to its final volume
with a prescribed diluent
Volumetric flask
o designed to hold different volumes rather than one exact amount
o Erlenmeyer flask has a wide bottom that gradually evolves into a smaller, short neck
o Griffin beaker has a flat bottom, straight sides, and an opening as wide as the flat base, with a small spout in the lip.
o Used for preparation of reagent
o Designed to hold different volumes rather than a single amount; has different graduations unlike
volumetric flask
Erlenmeyer flasks and Griffin beakers
o has calibration marks along its length and is used to measure volumes of liquids
o long, cylindrical tubes usually held upright by an octagonal or circular base
o do not have the accuracy of volumetric glassware it has calibration marks along its length
o used to measure different volumes of fluid which could vary from 10mL, 25 mL, 50 mL, etc.
Graduated cylinders
- instrument used to transfer liquids from one vessel to another
- Glass or plastic utensils used to transfer liquids; they may be reusable or disposable
- Designed to contain (TC) or to deliver (TD) a particular volume of liquid
PIPETS
PIPET DESIGN
To Contain (TC)
To Deliver (TD)
o holds or contains a particular volume but does not
dispense that exact volume
To Contain (TC)
will dispense the volume indicated
To Deliver (TD)
PIPET DRAINAGE CHARACTERISTICS
Blowout and Self-draining
o has a continuous etched ring or two small, close, continuous rings located near the top of the pipet
o the last drop of liquid should be expelled into the receiving container; exact volume is only obtained
when the last drop is blown out
Blowout
o the user allows the contents of the pipet to drain by gravity
Self-draining
Measuring or Graduated Pipets
PIPET TYPES
A. Mohr
B. Serologic:
C. Micropipet
does not have graduations to the tip
o self-draining pipet, but the tip should not be allowed to touch the vessel while the pipet is draining
o calibrated in between two marks
Mohr
has graduation marks to the tip\
o used for the measurements of reagents and are not generally considered accurate enough for measuring viscous samples and standards
o Blowout pipet
Serologic
with a total holding volume of less than 1 mL
- may be designed as either a Mohr or serologic
pipet
Micropipet
- Ostwald-Folin
- Volumetric
- Pasteur
- Automatic
Transfer Pipets
used with biologic fluids having a
viscosity greater than that of water
o blowout pipets, indicated by two etched continuous rings at the top
o ideal for transferring viscous solution
o has a bulb near the tip of the pipet
Ostwald-Folin
designed to dispense or transfer aqueous
solutions and is always self-draining
o has the greatest degree of accuracy and precision and should be used when diluting standards, calibrators, or quality-control material
o has a bulb at the middle of the pipet
o designed to deliver a fixed volume of aqueous solution
Volumetric
do not have calibration marks and are used to transfer solutions or biologic fluids without consideration of a specific volume
o not used in any quantitative analytic techniques
o disposable pipets
o may or may not have calibration marks
Pasteur
most routinely used pipet in today’s clinical chemistry laboratory
o Safe, stable, high precision, saves time, less cleaning required
Automatic
can aspirate and transfer only a single
volume
Fixed
different volumes however, only one
volume can be used at a time
Variable
relies on a piston for suction
creation to draw the sample into a disposable tip that must be changed after each use
Air Displacement
operates by moving the piston in the pipet tip or barrel, much like a hypodermic syringe
▪ the piston itself moves into the pipet tip, like
in syringe.
▪ Does not require a different tip for each time.
▪ There is a possible carry over at times
Positive Displacement:
obtain the liquid from a common
reservoir and dispense it repeatedly
▪ May be bottle top, motorized, handheld
Dispenser
- looks like a wide, long, graduated pipet
with a stopcock at one end - used to dispense a particular volume of
liquid during a titration - used in quantitative chemical analysis
used to measure the volume of a liquid
BURETS
- Sometimes used for transfer of small volumes in blood gas analysis or in separation techniques such as
chromatography or electrophoresis
o Used if < 500 µL of solution
- Glass and have fine barrels
SYRINGES
o Drying agents to make sure that your chemical is free from any moisture that may alter testing
o Placed below the perforated platform of a desiccator
o keep other chemicals from becoming hydrated
o most effective when placed in a closed, airtight chamber called a desiccator
o In the laboratory, desiccants are primarily used to prevent moisture absorption by chemicals, gases,
and instrument components.
Dessicants
required for the preparation of any
primary standards
o mechanical analytic balance is also known as a substitution balance
Analytical Balance
uses an electromagnetic force to
counterbalance the weighed sample’s mass
- measurements equal the accuracy and precision of any available mechanical balance, with the advantage of a fast response time
Electronic Balance
- Separates solid from a liquid suspension by means of a
centrifugal force - A process in which centrifugal force is used to separate
solid matter from a liquid suspension - Centrifugal force: mass, speed, and radius
o Speed: rpm
o Centrifugal force generated: RCF or g
𝑅𝐶𝐹 = 1.118 × 10−5 × 𝑟 × (𝑟𝑝𝑚)
2
Centrifugation
o daily cleaning of any spills or debris
o balancing the centrifuge load
o centrifuge cover should remain closed until the centrifuge has come to a complete stop
Centrifuge Care
sed to check the speed of
a centrifuge
o In using a centrifuge,
make sure that it is
covered from start to
finish to avoid aerosol
contamination
o Do not manually
stop a centrifuge.
Wait for it to stop
itself.
Tachometer or Strobe Ligh
- Used for the separation of solids from liquids
- Filter material is made of paper, cellulose and its derivatives, polyester fibers, glass, and a variety of resin column materials
- Filter papers differ in pore size
o Should be selected according to separation needs - Filter paper should not be used when using strong acids
or bases
Filtration
Liquid that passes through the filter paper
FILTRATE
- Used for separating macromolecules from a solvent or smaller substances
- Large particles from small particles
Dialysis
: fluid that passes
Dialysate
fluid that did not pass
Retentate
- Freeze drying
o Liquid to powder-like
Lyophilization
- separation of compounds, based on their relative
solubilities in two different immiscible liquids or solid
matter compound - 2 Types:
o Liquid-Liquid Extraction
o Solid-Liquid Extraction
Extraction
o Quantitative measurement of different chemical constituents of blood
Blood
o Checking of kidney functions
▪ Glomerular filtration rate
Urine
o Measurement of glucose and protein
CSF
SPECIMENS IN CLINICAL CHEMISTRY
- Blood
- Urine
- CSF
- Pleural fluid
- Pericardial fluid
- Peritoneal fluid
- Amniotic fluid
