KUHNES METHOD 99 



and dehydrating processes usually employed. For staining 

 Kiihne prefers a methylene blue solution prepared as 

 follows : 10 c.c. of a saturated alcoholic solution of methy- 

 lene blue is added to 100 c.c. of 5 per cent, solution of 

 carbolic acid (phenol). The sections are placed in this 

 solution for thirty minutes, then washed in water, and de- 

 colourised in very dilute hydrochloric acid (2 drops of strong 

 acid in 100 c.c. water). This decolourising operation must 

 be very carefully conducted, as very thin sections will only 

 require to be immersed for two or three seconds, after 

 which the sections are at once transferred to an alkaline 

 solution prepared as follows : 10 drops of a saturated solu- 

 tion of lithium carbonate in 10 c.c. of water. The sections 

 are then washed in water for a few minutes, dehydrated in 

 absolute alcohol, which Kiihne colours with a little methy- 

 lene blue. The sections are now placed in aniline oil, 

 which also contains a little dissolved methylene blue.* 

 The sections are next washed in colourless aniline, then in 

 xylol, and lastly mounted, as usual, in balsam. 



Ziehl-Neelsen Method. — This important method is used for 

 the diagnosis of tubercle and leprosy. These are the only 

 two organisms ordinarily met with which withstand the 

 decolourising action of the strong sulphuric acid used. The 

 following is the method employed for the examination of 

 tuberculous sputum and the bacilli of tubercle and leprosy 

 in sections. In the case of sputum, a very thin layer is 

 spread over the cover-glasses with a platinum wire or a 

 fragment of wood ; the cover-glasses are then dried and 

 fixed in the usual way. The cover-glasses are now floated, 

 the prepared side downwards, on a warm Ziehl's carbol- 

 fuchsine solution for ten minutes (three minutes in the 



* The aniline oil blue solution is prepared by shaking an excess of 

 methylene blue with colourless aniline, when, after standing with 

 frequent agitation, the coloured oil is filtered off. 



7—2 



