32 CLINICAL BACTERIOLOGY AND 



Mix an equal given quantity of the two solutions to- 

 gether, and flood the specimen already prepared with it. 

 Hold it over flame of spirit lamp until it nearly boils. Put 

 it aside for one minute, and wash in water j dry, stain with 

 aniline gentian violet for a second or two, wash in water, 

 dry, and mount. 



BonhilVs Method. — Take a small quantity of culture 

 material on a platinum loop, and move it about in a little 

 distilled water placed in a test-tube. Leave the tube un- 

 disturbed for six minutes, and then place 1 drop on a clean 

 grease-free cover-glass and dry in air. Now fix in the 

 flame, taking care not to overheat, and pour some orcein 

 solution on a watch-glass; float the cover-glass specimen 

 side downwards on the surface of the stain, and heat gently, 

 leaving the specimen in the stain for fifteen minutes. Wash 

 in water and examine, and if satisfactory mount in balsam, 

 but if not re-stain. 



Spore-Staining. 



The fact that spores are difficult to stain, while the rest 

 of the organism is not, suggests that the former is covered 

 by a specially thick envelope, which prevents the ingress 

 of the stain. 



Method I. — Take a film made in the ordinary way, and 

 flood with carbol-fuchsin and warm for fifteen to twenty 

 minutes. Wash in water, and dip for a second or so in 

 1 per cent, solution of sulphuric acid. Wash lightly, and 

 counter-stain by methylene blue ; wash, dry, and mount. 



Method II. — Moeller stain : 



Prepare slide in the ordinary manner. Keep in absolute 

 alcohol for two minutes, and then in chloroform. Wash in 

 water, allow the slide to stand in a 5 per cent, solution 

 of chromic acid for two minutes, wash, and then stain with 

 warm carbol-fuchsin for ten minutes. Wash, decolorize 

 in a 1 per cent, solution of sulphuric acid. Wash, counter- 

 stain with methylene blue for one minute. Wash, dry, and 

 mount. 



