CAPSULE STAINING. 31 



Neisser's stain for diphtheria bacilli. 



Solution No. i. Solution No. 2. 



Methylene blue, o.i gram. Bismarck brown, .2 



Alcohol, 2 c.c. Water (boiling), 100 c.c. 



Glacial acetic acid, 5 c.c. Dissolve the stain in the boiling 



Distilled water, 95 c.c. water and filter. 



Dissolve the methylene blue 

 in the alcohol and add it to the 

 acetic acid water mixture. Filter. 



To stain: Fix the preparation. Pour on the dilute acetic acid 

 methylene blue solution and allow to act from thirty to sixty seconds. 

 Wash. Then pour on the Bismarck brown solution, and after thirty 

 seconds wash off with water. Dry and mount. The bodies of the 

 bacilli are brown with dark blue dots at either end. 



Neisser recommends only five seconds as the time of application of 

 each solution. He also recommends that the culture be only nine to 

 eighteen hours old and that the temperature of the incubator do not 

 exceed 36 C. Incubation at 37 C. gives satisfactory results. 



Formulae for the Romanow r sky stains and for carbol thionin are 

 given in the section on blood. 



Capsule Staining. The best method for studying bacteria, as to 

 presence of capsules, is in the hanging drop, with the greater part of the 

 light shut off by the diaphragm. 



In material where capsules are w r ell developed, as in pneumonic 

 sputum, the Gram method of staining brings out the capsule perfectly. 

 This is of diagnostic value, as the more or less nonpathogenic pneu- 

 mococci common about the mouth do not seem to show a capsule 

 when stained in this way. 



The most beautiful method of staining capsules is the latest one 

 proposed by Muir. 



1. Prepare thin film,- dry and stain in carbol fuchsin one-half 

 minute; the preparation being gently heated (steamed). 



2. Wash slightly in 95% alcohol, then wash well afterward 

 in water. 



3. Flood preparation in mordant for five to ten seconds. 



