66 ANILIN COLOURS GIVING INDIRECT NUCLEAR STAINS. 



washing-out processes. Of course you will have to stain 

 longer in the weaker solutions. As to the anilin solutions, 

 see ante, 96. 



Washing out and Clearing. 



For general directions for washing out and clearing, see 

 above, 97 and 99. 



FLEM MING'S first method (1. c. in last ). Wash out with 

 pure alcohol, followed by clove oil. This method stains 

 resting chromatin as well as " mitoses." 



FLEMMING'S second method (Zeit.f. wiss. Mik., i, 3, 1884, p. 

 350). Wash out until hardly any more colour comes away, in 

 alcohol acidulated with about 0*5 per cent, of hydrochloric 

 acid, followed by pure alcohol and clove oil. (You may use 

 the HC1 in watery solution if you prefer it.) The strength 

 given appears unnecessarily high, and I therefore generally 

 use with good results an alcohol of about 0*2 per cent, of HC1.* 

 (Objects are supposed to have been well fixed twelve hours 

 at least iuihe strong chromo-aceto-osmic mixture, and stained 

 for some hours.) PODWYSSOZKI (Beitr. z. path. Anat. v. Ziegler 

 u. Neuwerk, i, 1886; Zeit. f. wiss. Mik., iii, 3, 1886, p. 405) 

 prefers to stain for half an hour only, and wash out with O'l 

 per cent of HC1 in alcohol. In each of these ways you get 

 "mitoses" and nucleoli alone stained (if the fixation have 

 been performed as above directed). 



FLEMMING'S orange-method (very important) is given in the 

 chapter on "Combination Stains" ( 258). 



PODWYSSOZKI (1. c.) gives another method, which consists in 

 washing out (for from a few seconds to two minutes) in a 

 strongly alcoholic solution of picric acid, followed by pure 

 alcohol. Same results (except that the stain will be brownish 

 instead of pure red). 



BABES employed for washing out after staining in the 

 aqueous or alcoholic solutions above mentioned, pure alcohol 

 followed by oil of turpentine. For sections stained in the 

 anilin solution he recommends treatment with iodine, accord- 

 ing to the method of GRAM (see what is said as to the process 



* In Flemming's latest work (on the achromatic structures of the cell) he 

 has been using a lower strength than this even, viz. O'l per cent, at most, see 

 Arch.f. mik. Anat., xxxvii, 1891, p. 249. 



