Blood Staining

Blood Staining

AIM

To make a blood smear ,stain it using Field’s stain and observe the erythrocytes and leucocytes.

Theory and Principle:

Blood smears are used to determine leukocyte differentials, to evaluate erythrocyte, platelet and leukocyte morphology, and, if necessary, to estimate platelet and leukocyte counts. It is also used for diagnosis of parasites like plasmodium in the blood.

Field’s Stain is a romanowsky stain, used for rapid processing of blood specimens and is used to stain thick and thin films. It consists of two differential stain.Field stain A which is methylene blue and Azure dissolved in a phosphate buffer solution.It is the basic component of the stain and Field stain B made up of Eosin Y in a buffer solution which is the acidic component of the stain.These basic and acidic dyes induce several colours when applied to cells.The fixator, methanol, does not allow any additional changes to the slide. The basic component of peripheral white blood cell( cytoplasm) is stained with acid dye and the acid component that is nucleic acid of the nucleus takes on the basic dye and is stained blue to violet. The neutral components of the cells are stained by both dyes(Field’s stain A and B solution).

Requirements

• Cotton

• Spirit

• Blood sample

• Clean grease free slides

• Methanol fixative

• Field’s stain A and Field’s stain B.

Procedure

1. Finger Prick under aseptic condition.

2. Place a small drop of blood, on one side about 1-2 cm from one end of a slide.

3. Without delay place another slide at an angle of 45° to make contact with the drop.

4. Spread it over an area of about 2 cm2(The film should be distributed so thinly that it appears transparent.

5. After air drying the thin blood film,immerse or fix the smear in methanol for 1 minutes.

 6. Flood or dip the slide in Field’s Stain A for 2-3 seconds.

7. Wash it with distilled water,

8. Flood or dip the slide in Field’s Stain B for 2-3 seconds and wash with distilled water.

9. Now air dry the smear and observe under microscope

Diagram

Observation

Results

The blood smear was stained using field’s stain and erythrocytes and leucocytes were observed under microscope.