164 COMBINATION STAINS. 



261. Methyl Green and Eosin (LIST, Zeit. f. wiss. Mik., ii, 1885, 

 p. 147). Stain for a few minutes in a mixture of three parts of absolute 

 alcohol with one part of aqueous solution of eosin (0'5 per cent.), wash, and 

 stain for five minutes in 0'5 per cent, aqueous solution of methyl green. 

 Wash, dehydrate, clear, and mount in balsam. The preparations do not keep 

 well in glycerin. 



The method may be varied (1. c., p. 150) by diluting the methyl green 

 solution with fifty volumes of water, and staining for twenty-four hours. 



It may also be varied by diluting the original methyl green solution with 

 three volumes of absolute alcohol. 



The preparations should not be left in the alcohol used for dehydrating 

 after the colour of the eosin has begun to reappear. 



261a. Methyl Green and Eosin (RHUMBLEE, Zool. Anzeig., No. 411, 

 1893, p. 47). Make a mixture of equal parts of 1 per cent, aqueous methyl 

 green solution, of a solution of 0'8 grm. of eosin in 50 per cent, alcohol 

 (quantity not stated), and of absolute alcohol. Stain (sections or small 

 pieces of material fixed with picro-sulphuric acid or alcohol) for half an hour ; 

 wash out first with water, then with successive alcohols, for not more than 

 a quarter of an hour, clear and mount. The novelty contained in this paper 

 consists in the somewhat astonishing statement that the stain serves to 

 differentiate living substance from dead or inorganic substance. " All sub- 

 stance that was living at the time of fixation being stained of a sharp red, 

 all dead substance or inorganic substance of a sharp green." 



262. Methyl Green and Bismarck Brown (LIST, Zeit. /. wiss. 



ii, 1885, p. 145). Stain for a few minutes in Weigert's Bismarck brown 

 ( 110), wash, and stain in 0'5 per cent, aqueous solution of methyl green. 

 Clear with bergamot oil or xylol, and mount in balsam. 



Or, dilute the Bismarck brown for staining with three volumes of absolute 

 alcohol, wash out with strong alcohol, and stain for a few minutes in the 

 methyl green solution diluted with three volumes of absolute alcohol. 



Or, stain for twenty-four hours in the Bismarck brown solution diluted 

 with fifty volumes of water, and then for twenty-four hours in the methyl 

 green solution diluted with fifty volumes of water. 



263. Methyl Green and Nitrate of Rosanilin (LIST, 1. c.). Stain 

 for a few minutes in 0'5 per cent, aqueous solution of methyl green, wash, 

 and stain (1. c., iii, 1886, p. 393) for ten to fifteen minutes in O'OOOl per 

 cent, aqueous solution of nitrate of rosanilin, wash out (rapidly) with abso- 

 lute alcohol, and clear. 



264. Anilin Green and Bismarck Brown (LiST, I. c.). To be used 

 in the same way as methyl green and Bismarck brown supra ( 262), and 

 giving much the same results. 



265. Anilin Green and Eosin (SCHIEFFEBDECKER, Arch.f. Mile. Anat., 

 xv, 1878, p. 30). Stain in a solution made by adding a few drops of aqueous 

 solution of eosin to a watch-glassful of alcohol for from half an hour to 

 several hours, wash with water, and stain for a few minutes in 1 per cent. 



