FLAG ELLA STAINING fi\ 



3. Wash well in water, and treat with 25 per cent, sulphuric 

 acid, or 33 per cent, nitric acid, until the film remains decolorised 

 when washed with water. 



4. Dry between layers of / If 

 fine filter paper, and counter- 



2 



G 



I 



stain with Loffler's alkaline 

 blue. 



5. Wash thoroughly, dry, and 

 examine. 



In the case of milk, the Fig. 8.-Heating stage, 



bacilli of tubercle or other acid- 

 fast organisms will be stained red, and the milk or casein cells 

 blue. 



FraenkePs modification of Ziehl-Neelsen method. — In this process the con- 

 trast stain is combined with the decolorising agent. The films are stained 

 as in the ordinary method, and then placed in a solution composed as follows : — 



Nitric acid . . . . . . .20 parts. 



Absolute alcohol . . . . . • 3© »» 



Distilled water . . . . . • 5° 1. 



Methylene-blue crystals to saturation . 



The stained films are treated with the above until the whole of the red 

 colour has been discharged and replaced by blue. 



Flagrella Staining* 



Successful staining of flagella is a question of practice, and of 

 careful and exact technique. Whateyer the method of staining 

 adopted, the preparation of the film is the same, and too much 

 care cannot be exercised at this stage. The slides should in no 

 case have been previously used, and they should be most carefully 

 cleaned in the manner described on p. 54. When taken out of 

 the alcohol, care should be taken not to touch the edges with the 

 fingers, and it is well to mark clearly one extreme corner of the 

 slide, by which it can be held between the forefinger and thumb 

 during the succeeding manipulations. The slide should be care- 

 fully dried and polished with a clean piece of old cambric, without 

 handling with the bare fingers. It should then be passed several 

 times through the flame, and set aside to cool. 



Preparation of the film. — i. The cultures for examination should 

 be upon agar, and should not be less than six, or more than 

 eighteen, hours old, if incubation has taken place at 37°. If incu- 

 bated at 22°, slightly older cultures may be employed. 



2. Inoculate with a trace of the youngest growth of the culture 



