Erythrocyte (Gr. erythros, red; kytos, cell) or red blood corpuscles are circular, anucleated, highly flexible, biconcave disc-shaped cells with high edges. The sixe of each cell averages 7.2 micrometer in diameter and 2.1 micrometer in thickness. It is 1.0 micrometer thick in the center. A complex membrane surrounds it, which is a bimolecular layer of protein. There is an inner most structure, called stroma, which is composed of lipids and proteins in the form of a fibrous protein. The cell contents are 90% hemoglobin. There are two methods for estimation of erythrocyte count:
- Manual or microscopic method
- Automated method
Hemocytometer with cover glass, compound microscope.
Hayem’s diluting solution is prepared as follows:
- HgCl2 0.05 gm
- NaSO4 2.5 gm
- NaCl 0.5 gm
- Distilled water 100 ml
EDTA anticoagulated venous blood or blood obtained by skin puncture is used.
- Wipe finger with cotton soaked with alcohol, with a sterile lancet do small prick on the finger tip. Use pipette. Aspirate blood to 0.5.
- Aspirate diluting Hayem’s solution to the 101 mark. It will give 1:200 dilution of the blood.
- Hold the pipette horizontally and role it with both hands between finger and thumb.
- Place the counting chamber, absolutely free from dust and grease, on the table and lay the cover glass in place over the ruled area.
- Discard the first two or three drops from the pipette. Charge the counting chamber by holding the pipette in an inclined position. Allow 3 minutes for the cells to settle.
- Locate the central square, which is divided into 25 medium sized squares. Each of the medium sized squares is further divided into 16 smallest squares.
- Count the erythrocytes in medium sized squares (80 smallest squares) using high power objective.
- In order to avoid confusion in counting, count all cells wihich touch the upper and left outer double line of the group of 16 squares as if they were inside the square. Neglect all those cells, which touch the lower and right inner line.
You may calculate total number of erythrocytes per cu mm of the blood as shown in the following.
Supose number of erythrocytes counted in 5 intermediate squares
= 1/5 sq mm
= 1/5 sqmm x 1/10 mm
= 1/50 cu mm
(1) Increased in numbers of RBC called polycythemia it is due to
• Bone marrow failure
• Erythropoietin deficiency (2ndry to kidney disease)
• Hemolysis (RBC destruction) from transfusion reaction
• Multiple myloma
• Nutritional deficiencies of (Iron, Copper, Folate, Vit B12, B6)
- Newborns 4.8-7.2 millions
- Children 3.8-5.5 millions
- Adult (Male) 4.6-6.0 millions
- Adult (Female) 4.2-5.0 millions
- Pregnancy slightly lower than normal