130 A MANUAL OF BACTERIOLOGY 



Flagella Staining 



Many organisms possess delicate protoplasmic pro- 

 cesses flagella in greater or less number ; but these 

 are not visible when the organism is examined in the 

 living condition (except by the use of dark-ground illumi- 

 nation), nor when the ordinary staining methods are 

 employed. In order to demonstrate them it is necessary 

 to make use of some special method, in which a mordant 

 is essential. 



For all methods of flagella staining the cover-glasses 

 or slides must be absolutely clean, the cultures recent, 

 and the growth sufficiently diluted to obtain the organisms 

 in an isolated condition. 



To clean slides. Rub the slides with a clean cloth and place on 

 a piece of clean wire gauze and heat with a smokeless flame for some 

 minutes (by this means grease is completely removed). Remove 

 the slides when cool, not before. Cover-glasses may be heated 

 in strong sulphuric acid and afterwards thoroughly washed in 

 distilled water. 



To make the suspension. A little of the surface growth is 

 removed with a platinum needle and transferred to 2-3 c.c. of 

 sterile tap water in a test-tube. The growth is diffused in the 

 water by shaking the needle to and fro, and not by emulsifying 

 on the side of the test-tube. The water should be at 37 C., and 

 the inoculated water tube should be placed in the warm incubator 

 for half an hour. A 1020 hour surface agar culture of the 

 organism is to be preferred, and growth only should be removed, 

 with as little of the culture medium as possible. 



To make the film. Several small drops of the inoculated water 

 are placed on a clean side or cover-glass with a small platinum 

 loop, and are allowed to dry in the air without spreading. 



(a) Shunk's method. As soon as the films are dry, they may 

 be flooded with a fixative solution (consisting of equal parts of 

 formalin and distilled water) which is allowed to act for two 

 minutes and then washed off with water. The mordant alone 

 will, however, usually suffice. 



