6 MICRO-ORGANISMS AND DISEASE. [CHAP. 



The method extensively and successively used for the demon- 

 stration and preservation of microscopic specimens of micro- 

 organisms in fluids, as blood, pus, and juices, is that of Weigert 

 and Koch, which consists in spreading out on a glass slide or 

 cover- glass a very thin film the thinner the better of the 

 fluid (artificial or natural culture medium), blood, pus, or juice, 

 and drying it rapidly by holding it for ten to twenty seconds 

 over the flame of a spirit-lamp or gas-burner. The most success- 

 ful preparations are obtained when the heating is carried on 

 for such a time that the film, having become opaque at first, 

 rapidly turns transparent. Several drops of the aniline dye to 

 be used are then poured over the specimen, and after remaining 

 on it from five to thirty minutes or more are poured off. The 

 aniline d} T es used are those that are commonly known as having 

 a great affinity for cell-nuclei (Hermann) ; they are also known 

 as the neutral or basic, but not the acid ones. The most useful 

 aniline dyes amongst them are those that are soluble in water ; 

 these are preferable to those soluble in alcohol, but in cer- 

 tain special instances (to be mentioned hereafter) some of them 

 are of definite use. Methyl-blue, methyl-violet, vesuvin, 

 Bismarck-brown, magenta, fuchsin, rosanilin, gentian- violet, 

 Spiller's purple, eosin, dahlia, purpurin, iodine-green, are the 

 aniline dyes commonly used. The washing is carried out in 

 all cases, except with tubercle-bacilli and the bacilli of 

 glanders, with distilled water, then with alcohol ; this latter 

 as a rule is to wash out the dye from all parts except the 

 micro-organisms, and it is therefore necessary not to carry the 

 washing further than this, but to carry it as far as practicable. 

 After this, wash again with distilled water, dry, and mount 

 in a drop of Canada-balsam solution. Throughout this pro- 

 cess it is of course necessary always to remember on which 

 side of the glass the specimen had been dried. 



Double staining is carried out with any two of the above 

 dyes ; as a rule a brown and blue or violet, or a red and blue, 

 are preferred. Some of the violet and purple dyes have the 

 peculiarity that in some not in all instances they give to 

 the preparation a double tint some things appear blue, 

 others more pink. 



The process of double staining is carried out either for 

 each dye separately i.e. we first apply one dye, after some 

 minutes wash in distilled water, and then apply the second 

 dye or the two dyes to be used are mixed and then used like 

 a single dye. 



In the case of tubercle-bacilli the staining is first done with 

 a magenta mixture (Ehrlich's, Weigert's, or Gibbes 3 see 



