CC PART 1 Flashcards
(2) WET CHEMISTRY
A. Liquid Reagent
B. Lyophilized Reagent
entails freezing a material at −40 °C or less and then subjecting it to a high vacuum. Very low temperatures cause the ice to sublimate to a vapor state. The solid non-sublimable material remains behind in a dried state.
Lyophilization
- In Jendrassik-Grof Method for bilirubin, caffeine and sodium benzoate serve as:
A. Color developer
B. Stopper of reaction
C. Dissociating reagent
D. Reducing agent
C. Dissociating reagent
Rationale:
● Technique for Multiple Choice Questions →
Elimination
○ Color developer (diazo reagent) ✖️
○ Stopper of the reaction (Ascorbic acid) ✖️
● Explanation: In Jendrassik Grof, B1 is attached to
the albumin. When you add a diazo, it will not
react. B1 and Albumin (Both are indirect method)
must be separated via Dissociating Reagent such
as
■ Caffeine
■ Sodium benzoate
■ Evelyn Malloy
● Reagent: 50% methanol
Storage and stability
Reagent storage:
2-80 C
- Performance indicator that refers to the dispersion of repeated measurements around the mean
a) Accuracy
b) Precision
c) Reliability
d) Linearity
B. Precision - magkakasama yung values pero wala sa gitna
Rationale:
● Accuracy: Nearness to the true value (If it hits the right target)
● Reliability: If the machine remains both accurate and precise for an extended period of time
● Linearity: It is the reportable range of an analyte
- Long acting barbiturate used in treating generalized grand-mal tonic-clonic seizures:
a) Ethosuximide
b) Acetaminophen
c) Tacrolimus
d) Phenobarbital
D. Phenobarbital
Rationale:
● 2 types of seizure:
○ Grand mal
○ Petit mal
● NOT tacrolimus since it is an immunosuppressant
● NOT acetaminophen since it is an antipyretic
- When the spread of data in the gaussian distribution falls between +/- 2 SD, the confidence
interval is:
a) 68.3%
b) 95.8%
c) 99.7%
d) 100%
b. 95.8%
- Convert 5 dL to mL
A. 5 mL
B. 50 mL
C. 500 mL
D. 5000 mL
C. 500 mL
Rationale: 1 dL = 100 mL
- Process that guarantees that the laboratory results are accurate as possible and includes three phases of testing: Preanalytic, analytic, post-analytic
A. Quality Management
B. Quality assurance
C. Quality points
D. Quality Control
Quality Assurance
Rationale: The bigger umbrella is Quality Management. Under Quality Management is the Quality Assurance. Quality Assurance has three phases called pre-examination,
examination, and post examination. Quality Control is under the examination phase or analytical phase.
- Which of the following lipoproteins can be observed in the upper part of the evacuated tube?
A. Cholesterol
B. VLDL
C. Triglycerides
D. Chylomicrons
D. Chylomicrons
Rationale:
● If the patient does not fast, this can be observed in the upper portion of Chylomicrons
● If the sample is turbid? = VLDL
What type of triglyceride does chylomicron transport?
● Exogenous
○ For VLDL
● Endogenous
If the question would be “Which LIPID…” = The answer would be Triglycerides
- Instrument technique that measures the amount of light emitted by the excited atom
a) Atomic Absorption Spectrophotometry
b) Spectrometry
c) Flame Emission Photometry
d) Nephelometry
Flame Emission Photometry
Rationale:
● If the question would be “The amount of light…”
○ Emitted: Flame Emission Photometry
○ Absorbed by ground state: Atomic
Absorption Spectrometry
○ Transmitted: Spectrometry
○ Scattered: Nephelometry
○ Blocked: Turbidimetry
- If the quality improvment, the question “Does this process need to exist?” refers to:
A. Lean
B. Six Sigma
C. Lean Six Sigma
D. Quality Assessment
A. Lean
Rationale:
● Lean: Reduction of waste
● Six Sigma: Reduction of errors
● If combined (Lean Six Sigma): Reduction of waste
and errors
- Sa blood sample collected by the medical technologist was accidentally left at the nurse station of hours, what test can still be performed?
A. Electrolytes
B. Cholesterol
C. Triglycerides
D. Glucose
C. Triglycerides
Rationale:
● Electrolytes: Increase
● Glucose: Decrease every hour
(2) CHEMISTRY APPROACH
1 WET CHEMISTRY
2 DRY CHEMISTRY
NO reconstitution needed
Liquid Reagent
Does Lyophilized Reagent Has Longer shelf life?
True or False
True
+ Diot H2oO -> Reconstituted
Lyophilized Reagent
Machine: Vitros
Dry Chemistry
Machine: Cobas
Wet Chemistry
(5) layers in Dry Chemistry
- Spreading
- Scavenger
- Reagent
- Indicator
- Support
Principle: Reflectance Photometry
- Lights are reflected
Dry Chemistry
Validation Studies
BUN STD = 10 mg/dL
a. 11 mg/dL
b. 10 mg/dL
c. 9 mg/dL
d. AOTA
b. 10 mg/dL (Accuracy)
Smaller volume requirement
Dry Chemistry
(5) Reagent Considerations
- Storage and stability
- Reagent Distribution
- Interfering Substances
- Expected Values
- Performance Characteristics
Change in the Reagent Lot number:
Perform Validation Study (Recalibration)
TREND or SHIFT
Using deteriorated reagent:
TREND
Trend and Shift is it Random or Systematic?
Systematic
(2) Reasons for using a calibrator/standard
- When performing method evaluation/technical validation
- Identification of errors in WG rule when the reagent is out of control
Must know of a medtech:
Knowledge of chemicals, standards solutions buffer and water requirements (MSDS/SDS)
(5) Reagent Deterioration
- Turbidity and precipitation
- Reagent fails to meet linearity claims
- Fails to recover control value in stated range
- Moisture has penetrated the vial and caking has occurred
- discrepancy in the first absorbance against water either greater or less than 0.8 depending on the set wavelength
Reportable range
linearity
(3) Interfering Substances
- Anticoagulants
- Drugs and other substances
- Pyridoxal Phosphate
(3) Common anticoagulants that cause interference:
- EDTA
- NaF
- Oxalate
(3) Blank
- Reagent blank
- Sample blank
- Water blank
If the sample is hemolyzed, lipemic, icteric what blank will you used?
Sample Black
To set spectrophotometer reading to zero
ABS -
% Transmission -
Water Blank
ABS - 0
% Transmission - 100
Before, During, After
Reagent
Before
Before, During, After
Sample
During
Before, During, After
Water Blank
Before
(4) Performance Characteristics
- Linearity/Analytical Measurement Range (AMR)
- Comparison of methods
- Sensitivity and Specificity
- Precision studies
Two commercial serum controls were assayed twenty times and the following ________________ precison was obtained.
Within Run
Two commercial serum controls were assayed for five consecutive days (triplicate for each level), the following ______________ precision was obtained
Run to Run Precision
Two means
T test
highly purified chemical than can be measured directly to produce a substance of exact known
concentration
Primary standard
substance of lower purity
Secondary standard
represents a specimen that is similar in composition to the patient’s whole blood or plasma. The value is known and measured the same way with the patient unknown sample.
Controls
Can you use Secondary standard alone?
No
(2) Controls
- Assayed
- Unassayed
High, Normal, Low
3 levels
Pathologic and nonpathologic
2 levels
All CC tests except Blood gases, therapeutic drugs, hormones
2 Levels
WG note:
1:2s
1:3s
R:4S
2:2S
4:1S
10:X
2 Levels
● Additional Westgard rules:
- 3:15
- 2 of 3:2S
- 7T
- 9X
3 Levels
Once in 24 hours
2 levels
Every 8 hours
3 levels
monitors overall reliability of laboratory results
means of accuracy and precision
Quality Controls
Two Divisions of Quality Control:
- Internal
- External
Proficiency testing
External quality control
- Analysis of control samples
- Instrument maintenance
- Statistical analysis
Internal quality control
Water purified by distillation, purified to remove almost all organic materials. Water is boiled, vaporized and condensed, distillation of water can be more than once.
Distilled Water
-process of removing dissolved gases by nonvolatile substances carried over steam in the distillation process or by dissolved substances from storage containers
Double Distillation
Water that is always used in Molecular biology
Double Distillation
used for tests that require Nh3 free water
Double Distillation
water purified by ion exchange ; removes some or almost all ions ;organic materials may still be present it is neither pure nor sterile ;deionized water is purified from previously pre-filtered or distilled water; produced using either an ANION or a CATION exchange resin
Deionized Water
water purified by reverse osmosis
Reverse-Osmosis
Uses pressure to pump water across a semipermeable membrane, thereby producing filtered product of the original water. Use also for pretreatment of water
Reverse-Osmosis
(4) Water Purification
- Activated charcoal
- Glass
- cotton
- submicron filter (0.2 mm)
Removes organic materials
Activated charcoal
Higher the water resistance/resistivity
_______ the water purity
Higher
used in clinical chemistry (highest purity)
Type 1
acceptable for most analytic requirements including reagent, quality control and standard preparation
Type 1
acceptable for most analytic requirements
Type 2
autoclave wash water- acceptable for glassware washing but not for analysis or reagent preparation
Type 3
- utensils made of glass or plastic that are used to transfer liquids. They may be reusable or disposable
Pipets
the tip should be immersed in the liquid to be transferred to a level that will allow it to remain in the solution after the volume of liquid has entered the pipet
Design
- lean by soaking to a soapy water with the pipet tips up; -used for volumes of 20 ml or less.
Pipets
- with different tolerances depending on the volume(mL).
Pipets
Holds a particular volume but does not dispense that exact volume; delivers the
amount by washing it and emptying
To contain (TC)
- Calibrated by: Mercury
- Drainage characteristic: Blow-out
- For viscous sample
To Contain
● Calibrated by: Water
● Drainage characteristic: Self-draining
● For non-viscous samples
To Contain
will dispense the volume indicated; transfer exact amount- do not blow the last drop; rate of flow is gravity
To deliver (TD)
(2) Drainage Characteristics
a. Blow-out
b. Self-draining
- Self draining
- Non viscous
To deliver (TD)
Markings are absent
Self-draining
Calibrated by: Distilled Water
To deliver (TD)
● Contains two etched rings with
colored rings near the top where
you place the rubber aspirator.
● Before, mouth pipetting was used but was discontinued a few years ago.
Blow-out
capable of dispensing several different volumes
Measuring or graduated
designed to dispense one volume without further subdivisions
Transfer
Serologic (graduated up to the tip)
Blow-out pipet
-self draining
- full volume should be used
-has dead space
Mohr (has dead space)
Pipets: According to function
- Measuring Pipets
- Mohr
- Serologic
a. Sahli-Hellige
b. Lang-Levy
c. RBC/WBC pipet
-do not have _____________
-transfer solution or biologic fluids without consideration of ________
-maybe disposable
-do not have CALIBRATION MARKS
-transfer solution or biologic fluids withOUT consideration of SPECIFIC VOLUME
-maybe disposable
-do not have _____________
-transfer solution or biologic fluids without consideration of ________
-maybe disposable
Pasteur Pipet
– for preparation of ____________________________
-always ______________.
- with greatest degree of accuracy and precision
- most accurate pipet
Volumetric pipet (bulb is in the middle)
– for preparation of ____________________________
-always ______________.
- with greatest degree of accuracy and precision
- most accurate pipet
- Non-viscous
– for preparation of STANDARDS/CALIBRATORS, CONTROLS AND REAGENT.
-always SELF-DRAINING.
- with greatest degree of accuracy and precision
- most accurate pipet
-with an etched marked near the delivery tip
-classified as blow-out pipette
- Viscous Samples
Ostwald Folin (bulb is nearer the mouthpiece)
-with an etched marked near the _______________
-classified as ___________________
-with an etched marked near the DELIVERY TIP
-classified as BLOW OUT PIPETTE
- Viscous Samples
- Single-Channel -
- Multi-Channel -
- Single-Channel - 1 Tip
- Multi-Channel - Multiple Tip
Fixed Volume and ___
Variable volume
for volumes less than 1 ml
-automatic implies that the mechanism that draws up liquid and dispenses it
Micropipet /Automatic pipet
- with total holding of less than 1ml
- may serologic or Mohr
Micropipet
QUESTION 1: Pipet with the greatest degree of accuracy and precision:
A. Serologic Pipet
B. Mohr Pipet
C. Ostwald-Folin pipet
D. Volumetric pipet
D. Volumetric pipet
§ Uses a suction aspirate & dispense sample through polypropylene tip. 1 or 2 stop. 2nd stop button is depressed to 2nd stop to “blow out”. tips can only be used once. Seals require periodic lubrication
Air Displacement
§ piston-mechanism; air movement; relies on a piston for suction creation to draw the sample into
a disposable tip.
Air Displacement
§ Uses a glass capillary tip fitted with Teflon -tipped plunger. No carryover. Tips are reusable. Plunger setting must be checked & teflon tip replaced periodically
Positive Displacement
§ Piston mechanism; syringe like; operates by moving the piston in the pipet tip or barrel; much like a hypodermic syringe.; piston comes in contact with the liquid; no air-interface
Positive Displacement
QUESTION 2: Positive displacement pipet:
A. With no air cushion
B. Can pipet high density samples
C. Aspiration force remains constant
D. A, B and C
D. A, B and C
- bottle-top
- motorized
- band-held
- attached to dilutor
Dispenser pipets
Obtain from a common reservoir and dispense it repeatedly
Dispenser pipets
for automatic pipetting devices
a. spectrophotometer – molar extinction coefficient of a compound is obtained or comparison of different dilution of potassium dichromate and its absorbance
b. Use potassium dichromate (K2CR207), a primary standard that is readily available and has a sharp absorption peak at 350 nm.
Photometric method
done by delivering and weighing a solution of known specific gravity; such as water; most desirable method-most desirable for pipet calibration.
Gravimetric method
molar extinction coefficient of a compound is obtained or comparison of different dilution of potassium dichromate and its absorbance
Spectrophotometer
A process whereby centrifugal force is used to separate solid matter from a liquid suspension
Centrifugation
Bottle: Low actinic
Positive Displacement
supernatant –
sediment –
supernatant – liquid
sediment – solid
Pipet Calibration
Quarterly or Every 3 months
parts include: head/rotor, carriers, shields, motor, chamber, control panel
Centrifugation
placing the tube by exact positioning or by the exact opposite
Rule of Thumb
PURPOSE OF THE HOLE IN centrifuge machine:________________________
For speed centrifugation
Relative Centrifugal Force (RCF): ________________
Revolutions Per Minute (RPM):________________
RCF: Force Acting in the Sample
RPM: Actual Speed
TWO MODELS OF CENTRIFUGE:
- Floor-standing centrifuges
- Bench-top centrifuges
Tube w/ gel separator:
Tube w/o gel separator:
**Tube w/ gel separator: 10-15 minutes @1000-3000g RCF
**Tube w/o gel separator: 10-15 minutes @1000
(2) How to obtain better separation of sample:
- Increase Speed
- Increase Time
(2) Influences the speed
- Motor
- Rotor
You can convert RPM to RCH
true or false
true
**Bouyant force
**Frictional force
Relative Centrifugal Force (RCF)
= 1.12X10^-5 x r x rpm2
RCF
(4) Types of Centrifuge
- REFRIGERATED CENTRIFUGE/ULTRACENTRIFUGE
- FIXED ANGLE
- SWINGING BUCKET
- CYTOCENTRIFUGE
OPERATION VERIFICATION:
- For critical applications, RPM is checked and recorded at least monthly
- For routine applications, RPM is checked and recorded at least quarterly.
The function of the refrigerated centrifuge is to protect the biological specimens from damaged caused by heating when centrifuge
Temperature ranges down to -10 C or -20 C (-50 F or -68 F)
Refrigerator
The cups are held in a rigid position at a fixed angle.
● This position makes the process of centrifuging more rapid than
horizontal-head centrifuge
● Less chance that the sediment will be disturbed when the centrifuge stop
Fixed Angle or Vertical Centrifuge
● Swinging bucket rotors have holders that swing outward in response to centrifugal force, so that the tube is held vertically when placed into or taken out of the holder, but held horizontally during centrifugation
SWINGING BUCKET / HORIZONTAL CENTRIFUGE
Characteristics of a Horizontal Centrifuge:
- When a rest, the tubes are in VERTICAL POSITION
- During centrifugation the tubes are in horizontal position
- Not capable of higher speed
QUESTION 3: Which of the following is not a characteristic of a Horizontal centrifuge?
A. Specimen cups are in vertical position when at rest
B. The cups move to a horizontal position during centrifugation
C. Capable of higher speed
D. As the sample is centrifuged, the particles being sedimented travel down through the liquid to the bottom of the tube
C. Capable of higher speed
QUESTION 4:
Which centrifuge is used when rapid centrifugation of solution is needed?
A. Horizontal-head.
B. Fixed-Angle
C. Cytocentrifuge
D. None of the items
B. Fixed-Angle
● Not used in clinical chemistry
● Used in histopathology
● Uses a very high-torque and low inertia motor to spread the
monolayers of cells rapidly across a special slide for clinical morphologic studies.
Cytocentrifuge
depending on usage: Clean interior components with soap and water followed by freshly made 10% v/v bleach solution, including sample buckets. Wearing protective gloves, wipe interior sides and bottom, taking care when removing broken pieces of glass
Weekly or biweekly
Place two equally balanced containers into the centrifuge, cover, and operate at the most commonly used speed, listening for unusual vibrations. Check the braking mechanism to ensure a smooth, gradual stop. Check the timer of the centrifuge at 15 minutes, 10 minutes, 5 minutes, and 1 minute for the time the centrifuge motor is spinning (reaches the desired rpm until the motor shuts off) using a stopwatch
Monthly
Inspect gasket and check for wear and defects, and inspect cover latch for appropriate seal. Inspect head, head shaft, and coupling for evidence of wear, cracks in fitting, corrosion, uneven wear, and signs of fatigue. Inspect brushes for wear and replace according to manufacturer’s
instructions.
Quarterly
Check the revolutions per minute at several commonly used speeds, including 3000 and 1500 rpm, while centrifuging a balanced load (after it has reached stable speed) using a tachometer aimed at the reflective strip viewed through the top of the centrifuge. If the revolutions per minute vary by more than 5%, consult the manufacturer’s instructions for solving the problem.
Quarterly
If an analog scale is provided for speed, the scale can be relabeled to match the actual speed. Lubricate the centrifuge shaft according to manufacturer’s instructions, if applicable.
Quarterly
- Used to separate serum from a blood clot-used in place of centrifugation
- Filter material is made of paper cellulose and its derivatives, polyester fibers, glass and a variety of resin column material
Filtration
- filter paper differs in terms of pore size and should be collected according to separation needs
- filter paper should not be used when using strong acids or bases
Filtration
- Filtrate : liquid that passes through the filter paper
Filtration
COLLIGATIVE PROPERTIES
4 properties based on the relative number of each kind of molecule present
- Vapor Pressure
- Freezing point
- Boiling point
- Osmotic pressure
(F and V)
(B and O)
Decrease (F and V)
Increase (B and O)
● As the freezing point decreases, the vapor pressure also decreases
● As the boiling point increases, the osmotic pressure also increases
Optically measures the level of color density corresponding to the amount of substance being tested for in the sample
Spectrophotometer
Sample is distributed evenly
Spreading Layer
Filters out substances that interfere with results
Scavenger Layer
Reagent reacts with sample
Reagent Layer
Reacted sample collects for spectral analysis
Indicator Layer
Optical interface
Support Layer
Pressure at which the liquid solvent is in equilibrium with the water vapor
Vapor pressure
Temperature at which the vapor
pressures of the solid and liquid
phases are the same
Freezing point
Temperature at which the vapor
pressure of the solvent reaches 1 atm.
Boiling point
What are the three methods of separation?
Centrifugation, Filtration, and
Dialysis
Pressure that allows solvent flow
through a semipermeable membrane to establish equilibrium between
compartments of different
concentrations.
Osmotic pressure
method for separating macromolecules from a solvent or smaller substances
Dialysis
For critical applications, RPM is checked and recorded
a. weekly or biweekly
b. at least monthly
c. at least quarterly
: at least monthly
What are the two operational verification activities for centrifuge set by CLSI?
- Check timer against standardized time source (stopwatch)
- Verification of speed
For routine applications, RPM is checked and recorded
a. weekly or biweekly
b. at least monthly
d. at least quarterly
d. at least quarterly
Verification of centrifuge’s speed is done using
using a tachometer or strobe
light
Weekly or biweekly cleaning uses soap and water followed by
: freshly made 10% v/v bleach
solution
Inspection of gasket and checking for wear and defects, and inspecting cover latch for appropriate seal is done
a. weekly
b. monthly
c. quarterly
c. quarterly
Checking the braking mechanism to ensure a smooth, gradual stop is done
a. weekly
b. monthly
c. quarterly
monthly
Checking the revolutions per minute is done
a. weekly
b. monthly
c. quarterly
c. quarterly
What do you call the product of filtration?
Filtrate
Checking the timer of centrifuge is done
a. weekly
b. monthly
c. quarterly
b. monthly
Used when rapid centrifugation of solutions containing small particle is needed up to 7000 rpm (slower than ultracentrifuge)
FIXED ANGLE / VERTICAL CENTRIFUGED
FIXED ANGLE / VERTICAL CENTRIFUGED
Angle:
25-40
