The hematologic stain that we use most frequently, and which was used in most of the plates pictured in this atlas, is Pappenheim's panoptic stain. It is based on a combination of the Jen-ner-May-Grunwald stain and Giemsa stain.
Method. Place the air-dried slide with the film side up in prepared May-Grunwald eosin-methylene blue solution for 3 min. Dilute with water or buffer solution (phosphate buffer pH 7.3, see below) for an additional 3 min. Pour off this solution and apply Giemsa stain immediately, without intermediate rinsing. The stock Giemsa stain is diluted with neutral distilled water by adding 10 mL water per 10 drops of Giemsa solution. Stain the specimen for 15 to 20 min. The dilution ratio and Giemsa staining time should be individually adjusted to allow for inevitable variations in the composition of the solution. After Giemsa staining, wash the slide with neutral water and tilt to air-dry. Fixation is effected by the methyl alcohol already contained in the May-Grunwald solution. The quality of the stain depends greatly on the pH of the water that is used. The smear will be too red if the water is too acidic and too blue if the water is too alkaline. Standard pH strips can be used to test the water for proper acidity. Water left standing in the laboratory can easily become too acidic through exposure to acid fumes, especially from carbon dioxide. The latter problem is solved by preboiling. A more accurate way to ensure correct acidity for staining is to use a pH 7.3 buffer solution (22.3 mL of 1/15 mol/L KH2PO4 + 77.7 mL of 1/ 15 mol/L Na2HPO4) instead of water.
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