Haemoglobin_Estimation_ with labelled microscope

Hemoglobin estimation through various methods and labelled microscope images

1. Haemoglobin Estimation Overview of Methods and Quality Control

2. Objectives To learn the various methods of haemoglobin estimation To have an overview of quality control in haemoglobin estimation

3. Introduction Haemoglobin (Hb) is the main component of RBCs Hb transports oxygen from lungs to tissues Contains 2α, 2β chains and 4 heme groups Hb iron content: 0.347 g/100 g Forms of Hb: OxyHb, CarboxyHb, SulphaHb, MetHb

4. Specimen for Hb Estimation Capillary blood from finger prick Intravenous sample with anticoagulant (EDTA)

5. Methods for Estimation of Hb Colorimetric methods Measurement of O₂ carrying capacity Measurement of iron content Specific gravity method

6. Colorimetric Methods Cyanmethaemoglobin method Oxyhaemoglobin method Electronic counter method Direct reading electronic haemoglobinometer Sahli’s method

7. Cyanmethaemoglobin Method Best method recommended by ICSH Principle: Hb + Drabkin’s fluid → Cyanmethaemoglobin Measured at 540 nm in spectrophotometer Advantages: No visual error, standard is stable Disadvantages: Turbidity & hyperbilirubinemia interfere

8. Oxyhaemoglobin Method Principle: Blood diluted in ammonia → OxyHb Measured at 625 nm Advantages: Quick, not affected by bilirubin Disadvantages: Standard not stable, misses SulphaHb

9. Electronic Counter Method Based on electrical impedance principle RBCs lysed, Hb converted to Cyanmethaemoglobin Concentration measured spectrophotometrically

10. Direct Reading Hb Meter Inbuilt filters and displays Hb concentration Works on cyanmetHb or oxyHb principle Disadvantage: Calibration errors possible

11. Sahli’s Method Principle: Hb + HCl → Acid haematin (brown) Compared with standard in colorimeter Advantages: Simple, cheap, bedside use Disadvantages: Visual error, comparator fades, unstable

12. Other Methods Alkali haematin method (NaOH) Haldane method (CarboxyHb with CO) Now rarely used

13. Normal Values of Hb Men: 13 – 18 g/dl Women: 12 – 16 g/dl Infants: 16.5 ± 3 g/dl

14. Errors in Hb Estimation Sampling error – venipuncture, anticoagulant Method error – varies with method used Equipment error – quality of material Operator error – improper training or overwork

15. Quality Control Essential for reliable results Internal quality control: repeat testing External quality control: cross-check with other labs Reference materials with known values are used

16. Take Home Message Hb estimation is vital for diagnosing anaemia Cyanmethaemoglobin is the gold standard method Electronic counters are widely used in labs Proper technique and quality control ensure accuracy