Abstract
Principle of method. Make a thick drop preparation of an accurately measured volume of marrow as in examination for malaria and count all the megakaryocytes in the preparation.
Method. From a well-shaken, evenly suspended marrow specimen, 1 fill a Sahli hemoglobin pipette to the 20 cram mark and blow contents onto a clean slide to form a thick drop. Spread evenly with the pipette tip over an oval area about 1.5 × 3 cm. Usually several such thick drop slides are made so that additional slides are available for examination in case part of the drop is detached by too vigorous washing and so that, when the count is low, a statistically significant number of cells can be counted. 1 Place slides in an incubator overnight. The following morning, immerse the slides in a solution of 5% formalin in 1% acetic acid in a Coplin jar until the erythrocytes are lysed and the thick drop is grayish in color. Flood the laking solution off the slide with water and gently rinse with buffer-phosphate solution (pH 6.4). 2 Stain with Wright's stain alone for 2 to 3 minutes. Add buffer-phosphate solution and stain 60 to 90 minutes. Wash slide by holding horizontally under gently running water. Air dry. Spread a thin film of immersion oil over the smear and examine systematically, using a mechanical stage and 200 X magnification, preferably with an 8 mm objective which is not immersed in the oil. Count all megakaryocytes seen in the specimen and multiply by 50 to express as number per ml. If the total nucleated marrow cell count per cmm and the leukocytic-erythrocytic ratio are determined, the number of megakaryocytes per million nucleated marrow cells or per million nucleated erythrocytic cells may be calculated.
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