COAL-TAB PLASMA STAINS. 2l7 



above, are liable to fade; by acidifying the mixture a 

 stronger and more sharply selective stain is obtained, which 

 does not fade. But too much acid must not be added, as 

 this would cause a staining of the iiiterfilar substances. The 

 following instructions for acidifying, due to M. HEIDENHAIN, 

 are from a paper of WARBURG'S (quoted from Zeit. f. >'/**. 

 Mil:., xi, 3, 1894, p. 383). To 2 c.c. of the Biondi mixture 

 (1 : 30) (by this is presumably meant the original mixture as 

 given above, but diluted with only 30 volumes of water 

 instead of 60) add 40 c.c. of distilled water, 3 c.c. of 0'5 

 per cent, solution of Saurefuchsin, and 0'2 c.c. of one fifth 

 per cent, solution of acetic acid (or, according to GROOVEN, 

 up. cit., xii, 3,, 1896, p. 379, four drops). [Grouven here 

 speaks of the mixture as " Triacid " (see next section) ; there 

 is a deplorable confusion in the nomenclature of these 

 stains.] 



Another process of acidification, complicated and difficult, is given by 

 M. HEIDENHAIN ( Veber Kern und Protoplasma, p. 116 ; Zeit.f. iciss. Mik., 

 ix, 2, 1892, p. 202) ; for this see last edition. See also ISRAEL (Praktilmm 

 Path. Hist., 2 Aufl., Berlin, 1893, p. 69) ; TBAMBUSTI (Ricerche Lab. Anat. 

 Roma, v, 1896, p. 82; Zeit. f. wiss. Mile., xiii, 1896, p. 357) ; and THOME 

 (op. cit. supra). 



After acidification the solution must not be filtered, for 

 filtration may render it less acid. If it has been kept for 

 some time a little more acid must be added ; for it will have 

 dissolved traces of glass, which is an alkaline body. M. 

 HEIDENHAIN therefore recommends that it be preserved in 

 rubber bottles. 



Before staining (M. HEIDENHAIN, loc. cit.), sections should 

 be treated for a couple of hours with 0*1 per cent, acetic 

 acid, then for ten to fifteen minutes with officinal tincture of 

 iodine, and be rinsed with alcohol before bringing into the 

 stain, in which they should remain for twelve to eighteen 

 hours. The treatment with acid is necessary in order to 

 ensure having the sections acid on mounting in balsam. The 

 primary object of the iodine is to remove any sublimate from 

 the preparations (Heidenhain's descriptions refer to sublimate 

 objects), but it also enhances the power of staining of the 

 chromatin with methyl green, and produces a more selective 

 staining of protoplasmic elements. 



