Trypanosomes and trypanosomiases / by A. Laveran and F. Mesnil ; tr. and much enl. by David Nabarro.
- Charles Louis Alphonse Laveran
- Date:
- 1907
Licence: Public Domain Mark
Credit: Trypanosomes and trypanosomiases / by A. Laveran and F. Mesnil ; tr. and much enl. by David Nabarro. Source: Wellcome Collection.
Provider: This material has been provided by the Francis A. Countway Library of Medicine, through the Medical Heritage Library. The original may be consulted at the Francis A. Countway Library of Medicine, Harvard Medical School.
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![less stain is necessary, and a cleaner preparation is ultimately obtained. Without previous fixation, run a few drops of stain on to the film, from a fine pipette (5 to 10 drops, according to the size of the film), and after thirty seconds run on double the number of drops of distilled water—again from a pipette—taking care that the water does not overstep the wax boundary lines. Tilt the slide carefully to mix the stain and the water, and allow the diluted stain to act for five to fifteen minutes. The stain is then poured away, the film is washed for ten or fifteen seconds in distilled water in a beaker, or blown on from a wash-bottle, and allowed to dry in the air. The slide may be preserved uncovered, or it may be covered in the ordinary way with a coverslip and xylol balsam, the latter, however, causing the stain to fade after a time.] [Marino's1 stain also gives very good results with trypanosomes. The alcoholic solution is obtained by dissolving in pure methyl alcohol the precipitate formed by adding an aqueous solution of eosin to an alkaline solution of methylene blue and azure blue in certain proportions.2 Four to 8 or 10 drops of the alcoholic stain are run on to the film (the number of drops varying with the size of the film), and left on for three minutes. Then double the number of drops of an aqueous solution of eosin (°'°5 gramme to 1,000 c.c. water) are added, and left on for two minutes, after which wash, dry, and mount. Some trypanosomes, such as those of birds and fishes, require rather longer to stain well—four to ten minutes with the blue, and eight to twenty minutes with the eosin.] If it be desired to stain a film rapidly, or if these special stains be not at hand, an alcoholic solution of fuchsin, or, better, a watery solution of magenta, or, better still, a solution of carbol thionin, may be used. Staining takes place rapidly, often in less than a minute. The nucleus, flagellum, and centrosome are stained more deeply than the protoplasm, and in this way sufficiently good results can be obtained when one wishes merely to ascertain the presence of the trypanosomes and their relative numbers. By Heidenhain's method (haematoxylin and iron alum) the nucleus, centrosome, and flagellum are stained darker than the protoplasm, but of course are less distinctive than when stained by our method or that of Romanowsky [or Leishman, Giemsa, etc.], in which they assume a tint different from that of the protoplasm. The methods of fixation and staining just described for blood will answer for other liquids which may contain trypanosomes, such as the blood-stained oedema fluids, cerebro-spinal fluid, culture media, anaemic blood, or blood diluted with salt solution or sodium citrate. In these cases, however, fixation is never so perfect as with pure blood ; often the trypanosomes in these liquids appear vacuo- 1 [Marino, Ann. Inst. Pasteur, v. 18, 1904, pp. 761-766.] 2 [The directions for preparing the stain are as follows : Take an aqueous solu- tion of methylene blue and azure blue (methylene blue, 05 gramme ; azure blue, 0-5 gramme ; water, 100 c.c.) and mix with an aqueous solution of sodium carbonate (0-5 per cent.). Keep the mixture in the 370 C. incubator for one to two days, or, better/in the thermostat at a higher temperature ; then add an aqueous solution of eosin to the blue mixture. The best strength of eosin solution varies with the quality of the blue, and must be ascertained by actually trying the effect of different strengths (o11 to 0*3 per cent.). The eosin blue mixture is then filtered, and the precipitate obtained is soluble in water and in methyl alcohol. The alcoholic solution mentioned in the text contains 0*04 gramme of the dried precipitate dis- solved in 20 c.c. pure methyl alcohol.]](https://iiif.wellcomecollection.org/image/b21172286_0035.jp2/full/800%2C/0/default.jpg)
