68 ANIL1N COLOURS GIVING INDIRECT NUCLEAR STAINS. 



Zells. Kern. u. Zellth., 1882, p. 384), and the stain may be 

 washed out with pure alcohol or (FLEMMING, Zeit. f. wiss. Mik., 

 1/1884, p. 350) with acidulated alcohol, as directed below for 

 safranin. But by far the best way of using it is, in general, 

 that due to BIZZOZERO (Zeit. f. wiss. Mik., iii, 1, 1886, p. 24). 

 The tissues may be hardened either in alcohol or in a chromic 

 mixture, but must in the latter case have been well washed 

 out with water. The staining solution is borrowed from that 

 of EHELICH for bacteria, and consists of 



Gentian violet ..... 1 part. 



Alcohol .... .15 parts. 



Anilin oil . . . . . . 3 



Water : . 80 



The sections are stained in it for five or ten minutes or 

 longer (for objects from Flernming's solution it will frequently 

 be advisable to stain for as many hours). After staining, 

 rinse the sections with alcohol, and bring them into a O'l per 

 cent, aqueous solution of chromic acid. After from thirty to 

 forty seconds bring them into alcohol, which begins the wash- 

 ing out of the colour. After thirty or forty seconds in the 

 alcohol put them back for thirty seconds into the chromic acid 

 (this is done in order to fix the colour more completely in the 

 nuclei) . Then bring them back into alcohol for thirty to forty 

 seconds, in order to wash out more colour and dehydrate them 

 at the same time. Then treat with clove oil, which will extract 

 more colour, and after a short time must be changed for fresh, 

 in which the sections remain until they are seen to give up 

 no more colour, when they are removed and mounted in 

 damar. 



You might give a longer treatment with alcohol, and a shorter treatment 

 with clove oil, but you would get a slightly different result. Alcohol washes 

 out colour freely from kinetic nuclei as well as from resting nuclei, whereas 

 clove oil acts much more energetically on the latter than on the former, and 

 thus serves to differentiate dividing nuclei. 



In some cases, especially those of tissues whose nuclei have 

 a tendency to give up the colour too freely, better results are 

 obtained by combining the foregoing method with that of 

 GRAM for the staining of bacteria (Fortschr. d. Medicin, ii, 1884, 

 No. 6; British Med. Journ., Sept. 6th, 1884, p. 486; Journ. 

 Roy. Mic. Soc. [N.S.], iv, 1884, p. 817). 



