MLPAO Laboratory Equipment and Quality Questions and Answers 2 — Questions and Answers

Question 1: What is the function of a micropipette calibration check using gravimetric analysis?

To verify the pipette delivers the correct volume by weighing dispensed water (Correct answer)
To check the pipette tip for contamination
To measure the viscosity of reagents
To verify the electrical calibration of the analyzer

Correct answer: To verify the pipette delivers the correct volume by weighing dispensed water

Gravimetric calibration checks a micropipette's accuracy by weighing aliquots of distilled water dispensed at the set volume, using the known density of water (1.00 g/mL at 20°C) to calculate the delivered volume.

Gravimetric calibration of micropipettes uses the relationship between water mass and volume (density of water ≈ 1.0000 g/mL at 20°C) to verify volumetric accuracy. The procedure: tare an analytical balance, dispense water from the pipette into a weighed container, record the mass, and calculate volume (V = mass/density). This is compared to the nominal volume. Acceptable accuracy is typically ±1-2% of nominal volume. Gravimetric checks are performed for laboratory pipette verification per quality management requirements. Micropipette calibration is essential because errors in pipetting volume directly affect reagent/sample ratios and thus test accuracy. ISO 8655 specifies performance standards for liquid-handling equipment used in Ontario laboratories.

Question 2: What is the purpose of the discriminator setting on a hematology analyzer for cell counting?

To set the temperature for cell lysis
To define voltage thresholds that distinguish cell populations by size or electrical impedance (Correct answer)
To calibrate the detector for specific cell types
To set the maximum cell count per analysis

Correct answer: To define voltage thresholds that distinguish cell populations by size or electrical impedance

Discriminators are electronic threshold settings on hematology analyzers that define boundaries between cell populations (e.g., between RBCs and platelets, or between WBC populations) based on electrical impedance or light scatter characteristics.

Modern hematology analyzers count and size cells using electrical impedance (Coulter principle) and/or light scatter (flow cytometry). As cells pass through an aperture or laser beam, they generate signals proportional to their size and complexity. Discriminators are electronic threshold voltage settings that define the boundaries between cell populations on histograms: the RBC/platelet discriminator separates platelets from RBCs; upper and lower WBC discriminators define the WBC counting region. These thresholds affect differential counts and cell population flags. Analyzers may automatically adjust discriminators based on histogram morphology, but technologists must review histograms for discriminator positioning when abnormal populations are present.

Question 3: Which of the following calibration materials is appropriate for calibrating a clinical chemistry analyzer?

Patient samples with previously known results
Commercially prepared calibrators with established analyte concentrations traceable to reference methods (Correct answer)
Quality control materials at normal concentration
Distilled water as a universal calibrator

Correct answer: Commercially prepared calibrators with established analyte concentrations traceable to reference methods

Calibrators are commercially prepared materials with certified analyte concentrations that are traceable to primary reference standards. They are used to set the analyzer's response curve, not QC materials.

Calibration establishes the mathematical relationship between instrument signal and analyte concentration. Proper calibrators must have: (1) Matrix matching (ideally human serum-based for serum assays); (2) Certified analyte values assigned by reference methods (e.g., IDMS for creatinine, reference spectrophotometry for enzymatic methods); (3) Traceability to SI units or NIST standards. Calibrators are distinct from QC materials — calibrators set the measurement function while QC monitors its stability. Using QC materials as calibrators is inappropriate. Calibration frequency varies by analyte and analyzer requirements. IQMH accreditation in Ontario requires documentation of calibration using appropriate materials.

Question 4: What is the purpose of preventive maintenance on laboratory analyzers?

To eliminate the need for quality control testing
To ensure consistent analyzer performance, extend instrument life, and prevent unexpected failures (Correct answer)
To recalibrate the analyzer after every patient run
To replace all consumable parts monthly regardless of need

Correct answer: To ensure consistent analyzer performance, extend instrument life, and prevent unexpected failures

Preventive maintenance (PM) involves scheduled cleaning, inspection, and replacement of components to maintain consistent analyzer performance, prevent unexpected breakdowns, and extend instrument lifespan.

Preventive maintenance programs for laboratory analyzers include daily, weekly, monthly, and annual tasks specified by the manufacturer and laboratory SOP. Tasks include: cleaning sample and reagent probes, replacing worn tubing, checking lamp intensity, verifying fluid levels, cleaning optical components, and replacing parts at defined intervals (e.g., lamp every 12 months, reagent probe seals monthly). PM documentation is required for laboratory accreditation. Consistent PM reduces carry-over, probe blockages, and spectrophotometer drift that can cause systematic QC failures and patient result errors. PM logs are reviewed during IQMH inspections in Ontario.

Question 5: An autoclave is used for sterilization in medical laboratories. What is the minimum recommended temperature, pressure, and time for a standard gravity cycle sterilization?

100°C, 0 psi, 10 minutes
121°C, 15 psi, 15-20 minutes (Correct answer)
134°C, 30 psi, 3 minutes
115°C, 10 psi, 30 minutes

Correct answer: 121°C, 15 psi, 15-20 minutes

Standard gravity displacement autoclave sterilization requires 121°C (250°F) at 15 psi steam pressure for a minimum of 15-20 minutes to achieve sterility (destruction of all microorganisms including resistant spores).

Steam sterilization (autoclaving) is the most widely used method for sterilizing heat-stable laboratory materials including glassware, media, biohazardous waste, and instruments. The standard gravity displacement cycle uses: 121°C (250°F) at 15 psi gauge pressure for 15-20 minutes contact time. This combination destroys all viable microorganisms including bacterial endospores (the most resistant form). The pre-vacuum cycle (132-135°C, 30-32 psi) is faster (3-4 minutes) and more effective for porous loads. Autoclave efficacy is verified using biological indicators (Geobacillus stearothermophilus spores) and chemical indicators. Proper loading is critical — bags must not be overfilled and must allow steam penetration.

Question 6: In flow cytometry, what does the parameter 'FSC' (forward scatter) primarily indicate about a cell?

Cell granularity or internal complexity
Cell size (volume) (Correct answer)
Surface antigen expression level
Cell viability (live/dead)

Correct answer: Cell size (volume)

Forward scatter (FSC) in flow cytometry is primarily related to cell size (volume). Larger cells scatter more laser light in the forward direction than smaller cells.

In flow cytometry, cells pass single-file through a laser beam and scatter light in different directions. Forward scatter (FSC) is detected at a small angle from the laser axis and correlates primarily with cell size/volume — larger cells scatter more light forward. Side scatter (SSC) is detected at 90° to the laser axis and reflects cellular granularity and internal complexity (nucleus size, cytoplasmic granules). On an FSC vs. SSC plot, different WBC populations cluster: lymphocytes (small, low FSC; low SSC), monocytes (larger, higher FSC; moderate SSC), and neutrophils (largest, highest FSC; high SSC due to granules). Flow cytometry is used in immunophenotyping leukemias and lymphomas in Ontario hematology laboratories.

MLPAO Laboratory Equipment and Quality Questions and Answers 2

What is the function of a micropipette calibration check using gravimetric analysis?