Laboratory Methodology in Hematopathology
Laboratory methods in hematopathology
Start with the preanalytical phase; most "hematology errors" are specimen problems, not assay problems. EDTA whole blood is the default for the CBC and morphology; citrate (3.2% sodium citrate) is used for coagulation and must be filled to the marked volume to preserve the 9:1 blood-to-anticoagulant ratio, since underfilling prolongs PT and aPTT. Heparin contamination from a line draw falsely prolongs clotting times and can distort smear staining.
Automated analyzers combine impedance (counting and volume), light scatter, and fluorescence (WBC differential, reticulocyte hemoglobin). They generate flags, not diagnoses: a "blast," "atypical lymphocyte," or "platelet clump" flag obliges smear review under predefined rules. On the smear, examine the monolayer near the feathered edge, estimate platelets (roughly 8–20 per oil-immersion field corresponds to 150–400 × 10^9/L), and correct the WBC count for nucleated RBCs. The 100-cell manual differential has wide sampling error; a 5% blast count could truly be anywhere near 1–12%.
Ancillary methods answer distinct questions: cytochemistry (MPO, Sudan black B, PAS, TRAP, Prussian blue iron), flow cytometry (CD antigens, kappa/lambda, aberrant maturation), karyotype and FISH, PCR/NGS (BCR-ABL1, JAK2 V617F, mutation panels), and hemoglobin analysis (HPLC, capillary electrophoresis, isoelectric focusing). Coagulation mixing studies (1:1 patient to normal plasma) separate factor deficiency (immediate correction) from inhibitor (no correction), with correction only after incubation suggesting a slow-acting inhibitor such as a lupus anticoagulant.
Trap: pseudothrombocytopenia from EDTA-induced platelet clumping - the analyzer reports severe thrombocytopenia while the smear shows platelet aggregates; repeat in citrate.
Rule of thumb: never act on a single unexpected automated result; verify with a smear or a repeat draw first.