Blood Smear: Preparation and Staining

 

Blood Smear: Preparation and Staining

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Reference:

1. Davidson, I. and Henry J., Clinical Diagnosis by Laboratory Methods, I. Davidsohn and J. Henry, eds., W.B. Saunders Co., Philadelphia 1974, pp. 135-137.
2. Wintrobe, M.W., et al., Clinical Hematology, Lea & Febiger,Philadelphia, 1974, p. 26.

Principle:

A thin blood smear is stained with Wright-Giemsa stain to enable evaluation of white blood cell, red cell and platelet morphology.

 

Specimen:

Peripheral blood or potassium EDTA anticoagulated blood (1 to 2 mg EDTA/1ml blood) may be used. Smears of peripheral blood must be made immediately. Smears may be made from EDTA-anticoagulated blood as long as two to three hours after collection. All specimens must be free of clots.

 

Procedure:

1. Anticoagulated specimens must be checked with two applicator sticks to be sure no clots are present. If clots are present the specimen is unsatisfactory.
2. A small drop of blood is placed on the surface of a clean glass slide near the end. If blood is taken from the finger, care must be taken to avoid touching the slide to the skin.
3. The slide is held between two fingers and the thumb of the left hand with the drip of blood on the upper surface towards the right. (Reverse for the left handed individual).
4. An edge of the spreader slide is placed on the first slide to the left of the drop of blood and is pulled to the edge of the drop. The angle between the two slides will vary according to the size of the drop and the viscosity of the blood. The larger the drop and the lower the hematocrit of the blood, the greater the angle must be to avoid running off the slide when spreading. Blood with a high hematocrit must be spread with a lower angle or the smear will be short and too thick to allow differentiation of cells. The approximate angle for normal blood is 30 to 40 degrees.
5. The drop of blood should be allowed to bank evenly behind the spreader which is then pushed to the left in a smooth, quick motion. The more rapid the motion, the shorter and thicker the smear. The smear should cover approximately half the slide with a gradual transition from thick to thin. No ridges should be present and the end (called the "feather edge") should be smooth and even. In the feather edge the red blood cells should not be routinely overlapped.
6. The smear is labeled on the frosted edge of the slide or the thick end of the blood with the patient's last name, first name, specimen number and date.
7. When the smear is completely dried, it may be stained by using the automated Hema-Tek II Slide Stainer. Automated Method:
   • The air dried smear is fixed with absolute methoanol and placed on a staining rack.
   • Allow slide to dry completely.
   • Prime stainer lines to remove air bubbles.
   • Place dried smear on stainer.

Reagents:

Hema-Tek II Stain Pak

 

Quality Control:

Strict adherence to procedure and visual check of the quality of stained slides.