COAL-TAR PLASMA STAINS. 227 



320. Artificial Alizarin (RAWITZ, Anat . Anz., xi, 10, 1895, p. 294). 

 RAWITZ has also worked out a process of obtaining a double stain 

 (chromatin and cytoplasm being stained of different colours) by means of 

 artificial Alizarin, or Alizarin -cyan in. The process is an adjective one, 

 requiring the use of special mordants supplied by the colour manufacturers 

 and is as follows : 



Both the Alizarin and the mordants are those prepared by the " Farb- 

 werke vorm. Meister Lucius & Briining in Hochst," and may be pro- 

 cured through Griibler & Hollborn. The Alizarin should be Alizarin 

 1, or SDG, or RX. A 5 per cent, suspension of it (it will not dissolve) 

 should be made in distilled water. The mordant should be that known as 

 CHROMBEIZE G A I or GA III. A stock solution should be made by mixing 

 70 parts of it with 130 of distilled water. 



Sections of Flemming material are brought into some of the stock 

 solution of mordant diluted with an equal volume of water ; sections from 

 chromic or chrome-picro-nitric material into the same diluted with 2 to 4 

 vols. of water ; sections from piero-nitric material into the same diluted 

 with 6 to 10 vols. of water. In either case they remain in the mordant for 

 twenty-four hours, and are then washed in distilled water until no more 

 colour comes away from them. 



They are then brought into the staining bath. This consists for 

 Flemming material of a portion of the stock suspension of Alizarin diluted 

 with 1 vol. of water ; for chromic and chrome-picro-nitric material of the 

 same diluted with 2 to 4 vols. of water; for picro-nitric material of the 

 same diluted with 6 to 10 vols. of water. To each staining bath there are 

 added a few drops of 1 per cent, solution of acetate of calcium (to be 

 procured from the Chemische Fabrik A. F. Kahlbaum, 35, Schlesischstrasse, 

 Berlin, S. 0.). They remain in the bath for twenty-four hours in the 

 warm, i.e. at a temperature of about 40 C. They are then washed for half 

 an hour to an hour in distilled water, then for one to two hours in 96 j>er 

 cent, alcohol, and are cleared with bergamot oil and mounted in balsam. 

 The sections must remain in the alcohol until all excess of Alizarin has been 

 removed from them, which may be known by their becoming clear and no 

 longer covered with a sort of fog. 



In the result, chromosomes ought to be deep reddish brown, plasma 

 orange. 



RAWITZ also gives, loc. cii., another process, with " Alizarin-cyanin RRR 

 doppelt," procured from the " Farbenfabriken vorm. Friedrich Bayer & 

 Co. in Elberfeld." Sections are mordanted for twenty-four hours in liquor 

 ferri sulfurici oxydati (see 254) diluted with 5 to 20 vols. of distilled 

 water, and are well washed with distilled water before coming into the 

 staining bath. This consists of a 5 per cent, suspension of the dye, diluted 

 with water just as for Alizarin, and with acetate of calcium added to it in 

 the same way. They are stained therein and further treated in all respects 

 as directed for Alizarin. The stain is a blue one. 



