Primary Method for Determining Hemoglobin Concentration on Automated Cell Counters
The primary method for determining hemoglobin concentration on most automated cell counters is a modified cyanmethemoglobin method (option D).
Measurement Principles in Automated Hematology Analyzers
Modern automated hematology analyzers employ spectrophotometric methods to directly measure hemoglobin concentration. The specific methodology has evolved over time, but remains based on the principles of the cyanmethemoglobin method with modifications for automation:
Modified Cyanmethemoglobin Method
- The traditional cyanmethemoglobin method has been modified to work in automated systems where reaction time is limited 1
- This involves:
- Rapid red blood cell lysis
- Conversion of hemoglobin to a stable compound
- Spectrophotometric measurement at specific wavelengths
- Direct quantification of hemoglobin concentration
Alternative Methods
While the modified cyanmethemoglobin method remains standard, some analyzers use variations:
- Sodium Dodecyl Sulfate (SDS) Method: Some analyzers use SDS to convert hemoglobin to a stable denatured globin hemichrome without cyanide 2
- Hemichrome Method: Some manufacturers use denatured globin hemichrome as the end product while still including cyanide in their reagents 3
Why Other Options Are Incorrect
Option A (Calculated by dividing Hematocrit by 3): This is an indirect estimation method sometimes used in clinical settings but is not the primary method in automated analyzers. Modern analyzers directly measure hemoglobin rather than calculating it 4, 5
Option B (Derivation from histogram): While histograms are used for cell counting and sizing in automated analyzers, they are not the primary method for hemoglobin determination 4
Option C (Laser light scatter): Light scatter technology is primarily used for cell counting and differentiation in automated analyzers, not for hemoglobin measurement 4
Clinical Implications
Accurate hemoglobin measurement is critical for:
- Diagnosing and monitoring anemia
- Guiding transfusion decisions
- Evaluating response to therapy
The College of American Pathologists and Clinical Laboratory Improvement Amendments (CLIA) set evaluation criteria of ±7% for acceptable hemoglobin measurement performance compared to reference methods 1.
Common Pitfalls in Hemoglobin Measurement
- Sample Integrity: Hemolyzed or lipemic samples can affect accuracy
- Calibration Issues: Improper calibration of analyzers can lead to systematic errors
- Interfering Substances: Certain medications or conditions can interfere with spectrophotometric measurements
- Sample Collection Method: Capillary fingerprick blood usually produces higher hemoglobin concentrations compared to venous blood 1
For optimal accuracy, laboratories should use only hemoglobin assay methods that are certified by the National Glycohemoglobin Standardization Program (NGSP) as traceable to reference standards 6.