Hemoglobin Estimation..clinical pathology

Hemoglobin is a protein present in RBCs for exchange of gases with tissues.  Blood is a specialized type of connective tissue in the form of fluid. It has two components called as plasma and cells.

• Plasma contains 12 coagulation factors. Serum is formed by the removal of these coagulation factors. Serum is collected in red color vacuotainer. It does not let the blood clot.
• The ratio of cells in the blood is RBCs, WBCs and Platelets as 500:1:30 respectively.

There are four methods for the determination of hemoglobin in the blood;

1. Sahli’s method..Outdated
2. Cyanmet Hb method..Updated
3. Spectrophotometer
4. Haden Haussen method

Sahli’s Method of Hemoglobin Determination:

Apparatus:

• Sahli’s tube which is having red and yellow scales on two sides. Red scale is percentage scale and yellow scale is gram percentage or g/100ml scale.
• Heamometer which is having two standards.
• Sahli’s pipette.
• Error percentage is 3%.

Procedure:

• In sahli’s tube, take N/10 HCL(1/10^th of the original HCL) up to 10^th level of scale.
• In sahli’s pipette, take 0.02ml(20microleter) blood.
• Add blood from pipette into tube.
• HCL will cause the lysis of the blood cells and hemoglobin is released.
• Hb after combining with HCL, forms acid hematin which is of tan color.
• Put tube in the hemometer and continuously add drops of distilled water and shake with the stirrer until color matches. Then, take the reading.

Interpretation of the results:

Heam portion of hemoglobin is a porphyrin type pigment and contains iron.

Types of Hemoglobin:

Heam Cycle: