20 MICROSCOPIC EXAMINATION OF BACTERIA. 
(f) Wash in 95 per cent. alcohol until no more 
color comes away. 
(g) Dryeand contrast-stain in safranin' (thirty 
seconds), or Bismarck-brown? (two to three min- 
utes), or eosin® (one minute). 
(2) Wash, dry, and mount. 
3. Staining Bacteria in Tissues. 
(1) First Method. 
(a) Harden the tissue in absolute alcohol or Zen- 
ker’s fluid.* 
(6) Dehydrate, embed, and section by the usual 
methods. 
(c) Stain as directed for cover-glass preparations, 
but a little more deeply. 
(d@) Draw the color with dilute acetic acid 
(1 : 1000) until the bacteria alone are stained. 
(e) Contrast-stain in eosin or any stain not re- 
quiring acid for differentiation. 
(f) Clear and mount. 
(2) Second Method. 
Stain the sections by Gram’s method as follows : 
(2) Stain in anilin gentian-violet for five minutes. 
(4) Wash in water. 
(c) Treat with Gram’s solution for one minute. 
(2) Wash in 95 per cent. alcohol until no more 
color comes away. 
1 Stock solution is ar per cent. solution of safranin in 
equal parts of methylated spirit and water. For use, dilute 
with 5 parts of water. 
* Saturated solution in equal parts of alcohol and water. 
5 Aqueous solution, I : 1000. 
4 See Appendix, page 173. 
