Routine Techniques

• What is the most common anticoagulant for routine hematology testing?

  EDTA (usually as K2EDTA)
• How soon should specimens for CBC be analyzed if stored at room temperature?

  Within 6 hours
• What is the maximum time specimens for CBC can be stored at 4°C?

  24 hours
• Why should PBS be prepared within 3 hours of collection?

  To reduce cell deterioration and artifacts
• What are the important components of specimen collection?

  • Equipment used to obtain specimen
  • Site and technique for venipuncture or skin puncture
  • Specimen handling after collection
• Why is it important to learn manual cell counts despite current technology?

  To understand fundamental hematology principles
• What equipment is used in manual cell counts?

  • Hemacytometer/counting chamber
  • Manual dilutions with calibrated automated pipettes
  • Diluents
• What is the principle behind manual cell counts?

  Same for RBC, WBC, and platelet counts
• What is the most commonly used counting chamber?

  • Levy chamber with improved Neubauer ruling
• What is the total area of the Levy chamber?

  3 mm x 3 mm
• How many squares are in each of the four WBC squares of the Levy chamber?

  Each is subdivided into 16 squares
• What is the area of each smaller square in the center square of the Levy chamber?

  0.04 mm²
• What is the general formula for manual cell counts?

  Total count =

  $\frac{cells\ counted \times dilution\ factor}{area\ mm^2 \times depth\ (0.1)}$
• How is 1 mm³ related to microliters?

  1 mm³ is equivalent to 1 microliter
• How do you convert microliters to liters?

  Multiply by a factor of $10^6$
• What is the most common dilution factor for erythrocyte counts?

  1:100 using isotonic saline
• What is the adult reference range for erythrocytes?

  • M: 4.2-6.0 x 10²/L
  • F: 3.6-5.6 x 10²/L
• What is the dilution factor for leukocyte counts?

  1:20 using a weak acid
• What is the adult reference range for leukocytes?

  3.6-10.6 x 10³/uL
• What is the dilution factor for platelet counts?

  1:100 using ammonium oxalate
• What is the adult reference range for platelets?

  150-450 x 10³/uL
• What diluting fluid is used for leukocyte counts?

  Weak acid solution
• What is the objective lens used for platelet counts?

  40x (LPO)
• What should be ensured for accuracy in manual cell counts?

  • Even distribution of cells
  • No more than 10 cell variation
  • Repeat count on the other side
• What is the reference range for leukocytosis?

  Greater than 11 x 10⁹/L
• What dilution factor is used if WBC count is above 30.0 x 10⁹/L?

  1:100 or 1:101
• What is the corrected WBC count formula?

  Corrected WBC count = $\frac{Uncorrected\ WBC\ count \times 100}{Number\ of\ NRBCs + 100}$
• What should be done if fewer than 50 platelets are counted on each side?

  Use a dilution of 1:20
• What should be done if more than 500 platelets are counted on each side?

  Use a dilution of 1:200
• How should platelet estimates be reported?

  • 0-49,000/uL: Marked decreased
  • 50,000-99,000/uL: Moderate decreased
  • 100,000-149,000/uL: Slight decrease
  • 150,000-199,000/uL: Low normal
  • 200,000-400,000/uL: Normal
  • 401,000-599,000/uL: Slight increase
  • 600,000-800,000/uL: Moderate increase
  • >800,000/uL: Marked increase
• What is the effect of EDTA on platelet clumping?

  It prevents platelet clumping
• What happens to mean platelet volume during the first hour in the tube?

  It increases
• What is a source of error in platelet counts according to Rodak?

  • Counting small RBC squares if normal plt count
  • Platelet satellitosis with EDTA
• What should be done if platelet satellitosis occurs with EDTA?

  Use sodium citrate and multiply by 1.1
• What is the reference range for platelets according to Rodak?

  150-450 x 10³/uL
• What is the reference range for platelets according to Brown?

  150-400 x 10³/uL
• What is the dilution factor for platelet counts if fewer than 50 platelets are counted?

  1:20
• What is the dilution factor for platelet counts if more than 500 platelets are counted?

  1:200
• What is the procedure if the difference in counts between sides of the chamber exceeds 10%?

  Repeat the counting procedure
• What is the significance of the Rule of Three in hematology?

  It helps verify the accuracy of blood counts