u 4 MICROSCOPIC METHODS. 



FraenkeVs modification of the Ziehl-Neehen Stain. 



Here the process is shortened by using a mixture 

 containing both the decolorising agent and the contrast 

 stain. 



The sections or films are stained with the carbol-fuchsin as above 

 described, and then placed in the following solution : 



Distilled water . . . . -5 parts 

 Absolute alcohol . . . .30 parts 



Nitric acid . . . . .20 parts 



Methylene-blue in crystals to saturation. 



They are treated with this till the red colour has quite disappeared 

 and been replaced by blue. The subsequent stages are the same as in 

 No. 5, supra. 



Leprosy bacilli are stained in the same way, but are 

 rather more easily decolorised than tubercle bacilli, and 

 it is better to use only 5 per cent sulphuric acid in 

 decolorising. 



In the case of specimens stained either by the original 

 Ziehl-Neelsen method, or by Fraenkel's modification, the 

 tubercle or leprosy bacilli ought to be bright-red, and the 

 tissue blue or brown, according to the contrast stain used. 

 Other bacteria which may be present are also coloured 

 with the contrast stain. 



The Staining of Spores. If bacilli containing spores are 

 stained with a watery solution of a basic aniline dye the 

 spores remain unstained. The spores either take up the 

 stain less readily than the protoplasm of the bacilli or 

 they have a resisting envelope which prevents the stain 

 penetrating to the protoplasm. Like the tubercle bacilli, 

 when once stained they retain the colour with consider- 

 able tenacity. The following is the method for staining 

 spores : 



1. Stain cover-glass films as for tubercle bacilli. 



2. Decolorise with I per cent sulphuric acid in water or with 

 methylated spirit. This removes the stain from the bacilli. 



