Page:Tropical Diseases.djvu/72

a minute; wash in tap water and then in distilled water; dry and mount in xylol balsam. This is an easily carried out and effective method of obtaining less intensely stained but more permanent preparations.

(b) The many methods of obtaining differential staining of protoplasm and nucleus depend on the circumstance that, when solutions of certain kinds of methylene blue and eosin are mixed, a third red dye, with a special affinity for chromatin, is formed. They are all modifications of the original Romanowsky method. They are a little uncertain and troublesome to use, their success depending on the purity of the chemicals employed and careful attention to detail. The beginner is advised to become proficient, in the first instance, with the simpler methods given above. Having attained this, he should practise some form of the Romanowsky method, for, besides displaying the intimate structure of the malaria parasite, in consequence of the intense tinting of the nucleus which it effects it greatly facilitates the finding of the smaller forms of the parasite, not always an easy matter, especially in the case of the very minute young subtertian or malignant parasite. Of the three methods given below, Leishman's, everything considered, is the best. Both it and Jenner's method have the advantage of dispensing with preliminary fixing.

Romanowsky's method.—Cover-glass films are fixed by heat 110° C. for one hour. They are then immediately floated for over two hours on a freshly prepared mixture of saturated watery solution of methylene blue, 2 parts, and watery solution of eosin (1 per cent.), 5 parts, washed in water, dried and mounted. The precipitate formed by the mixture must not be filtered out. Jennets stain.—Special care must be taken that there is no trace of alkali or acid on the glass on which the film is spread; it must, therefore, be washed in distilled water and stood in absolute alcohol. The stain is made by dissolving Grübler's water-soluble eosin (yellow shade) and medicinal methylene blue in pure absolute methylic alcohol, which must be free from acetone and other impurities; the solutions are then mixed in the proportions of 125 parts of 1/2 per-cent. solution of the eosin, and 100 parts of 1/2 per-cent. solution of the methylene blue. Or the compound body formed by the eosin and methylene blue in this mixture may be purchased in the dry state and subsequently dissolved in absolute methylic alcohol.