V. STAINING METHODS. BACTERIA STAINS. IN the following pages are collected the various "stock" stains in everyday use in the bacteriological laboratory, together with a selection of the most convenient and generally useful staining methods for demonstrating particular structures or differentiating groups of bacteria. The stains employed should either be those prepared by Grübler, of Leipzig, or Merck, of Darmstadt. The methods printed in ordinary type are those which a long experience has shown to be the most reliable, and to give the best results Methylene-blue.— 1. Saturated Aqueous Solution. Weigh out Methylene-blue 1.5 grammes Place in a stoppered bottle having a capacity of from 150 to 200 c.c. and add Allow the water to remain in contact with the dye for two weeks, shaking the contents of the bottle vigourously for a few moments every day. Filter. 2. Saturated Alcoholic Solution. Weigh out Methylene-blue. 1.5 grammes Place in a stoppered bottle of 150 c.c. capacity and add Alcohol, 90 per cent. 100.0 C.C. Allow the alcohol to remain in contact with the dye for two hours, shaking vigourously every few minutes. Filter. 4. Alkaline Methylene-blue (Löffler). Methylene-blue, saturated alcoholic solution. . 30.0 c.c. 100.0 " 1.5 grammes and proceed as in preparing the corresponding solution of methylene-blue. 10. Carbolic Gentian Violet (Nicollé). Measure out and mix Gentian violet, saturated alcoholic solution Carbolic acid, I per cent. aqueous solution Filter. 10.0 C.C. 100.0 66 Thionine Blue (or Lauth's Violet).11. Carbolic Thionine Blue (Nicollé). Weigh out Before use dilute with equal quantity of distilled water and again filter. CONTRAST STAINS. Eosin. There are several commercial varieties of eosin, which, from the bacteriological point of view, possess very different values. Grübler lists four varieties, of which two only are useful for bacteriological work: Eosin, aqueous yellowish. Lithium carbonate, cold saturated solution.. . 100.0 C.C. Filter. Picrocarmine. Weigh out Picrocarmine and dissolve in Distilled water . 2.0 grammes 100.0 C.C. 3. Mix 1 and 2, allow the mixture to stand fortyeight hours, then filter. 4. Add Glycerine Acetic acid, glacial 85.0 c.c. 10.0" 5. Allow the stain to stand for one month; then filter ready for use. Aniline Gentian Violet (For Weigert's Fibrin Stain).— Weigh out and place in a glass beaker. Measure out in a measuring cylinder, Distilled water 2.5 grammes 1.0 gramme 100.0 C.C. Place the beaker on a sand-bath, add the water in successive small quantities, and keep the mixture boiling for twenty minutes. Measure the solution and make up to 100 c.c. by the addition of distilled water. Filter. |