466 SOME ZOOLOGICAL METHODS. 



861. Stains forFlagella. The celebrated method of LOFFLEE 

 has run through several forms (Centralb. f. Bacterial., vi, 

 1889, p. 209 ; vii, 1890, p. 625; Zeit. f. wiss. Mik., vi, 3, 1889, 

 p. 359; vii, 3, 1890, p. 368; Journ. Roy. Mic. Soc., 1889, 

 p. 711; 1890, p. 678), of which that given here is the latest. 

 To 10 c.c, of a 20 per cent, solution of tannin are added 5 c.c. 

 of cold saturated solution of ferrous sulphate and 1 c.c. of 

 (either aqueous or alcoholic) solution of f uchsin, methyl violet, 

 or " Wollschwarz." The mixture will require for some forms 

 the addition of a few drops of 1 per cent, solution of caustic 

 soda; e. g. for typhoid bacilli, 1 c.c. ; for Bacillus subtilis, 28 

 to 30 drops; for bacilli of malignant oedema, 36 to 37 drops. 

 Some other forms will require besides the addition of a trace 

 of sulphuric acid to the soda solution, so for cholera bacteria, 

 half a drop to 1 drop ; for Spirillum rubrum, 9 drops. 



Cover-glass preparations are made and fixed in a flame in 

 the usual way, special care being taken not to over-heat. 

 Whilst still warm the preparation is treated with mordant 

 (i. e. the above-described mixture), and is heated for half a 

 minute until the liquid begins to vaporise, after which it is 

 washed in distilled water and then in alcohol. It is then 

 treated in a similar manner with the stain, which consists of 

 a saturated solution of fuchsin in anilin water, the solution 

 being preferably neutralised to the point of precipitation by 

 cautious addition of 0*1 per cent, soda solution. 



TEENKMANN (Centralb., vi, 1889, p, 433; Zeit. f. wiss. Mik., 

 vii, 1, 1890, p. 79) mordants for several hours at the normal 

 temperature in a 1 per cent, solution of tannin in 0*5 per cent, 

 hydrochloric acid, and stains for several hours in carbolic 

 fuchsin ; and gives also two other similar methods. 



BEOWN (The Observer, iii, 1892, p. 298; Journ. Roy. Mic. 

 Soc., 1893, p. 268) mordants for several hours in a mixture of 

 30 gr. tannin, 12 drops anilin oil, and 1 fl. oz, of alcohol, 

 which may, if required, be alkalised by addition of a trace of 

 caustic soda (so for Spirillum undula and Bacillus ulna), or 

 may be acidified for others with a little hydrochloric acid. 

 The cover is stained by the process of heating over a flame 

 for a few minutes with any anilin-water solution of fuchsin, 

 methyl violet, dahlia, methyl green, &c., neutralised with 

 caustic soda as in Loeffler's process, or with a solution of 

 rosanilin in anilin water. 



