52 EXAMINATION AND STAINING OF BACTERIA. 



with the mordant applied steaming, but not boiling, for three 

 minutes ; wash off thoroughly in water and dry ; treat with 

 the stain in the same manner as with the mordant ; wash in 

 water, dry, mount, and examine with an oil-immersion lens. 

 Other methods, such as Van Ermengem's, BowhilTs, etc., are 

 highly recommended, but in the author's hands have given 

 no better result than the foregoing three methods. 



VIII. The Staining of Bacteria in Tissues. 

 1. The Staining of Sections. 



Sections should be cut in the ordinary way in paraffin or 

 celloidin. The sections are first put into water for a few 

 minutes, then transferred to watch-glasses containing watery 

 solutions of any basic anilin dye, and allowed to remain from 

 five to ten minutes ; they are next removed, rinsed in water, 

 decolorized in a 0.1 solution of acetic acid for a few seconds, 

 again washed in water, then for a few minutes in absolute 

 alcohol, and placed in cedar oil or xylol. They are allowed 

 to remain in xylol from one-half to one minute. They, are 

 finally spread thinly on a spatula and brought to the slides, 

 where the excess of fluid is taken up with blotting-paper ; 

 after which a drop of xylol balsam is placed on the sections, 

 which are covered by thin clean cover-glasses, when they are 

 ready for examination. 



2. Gram's Method for Staining Bacteria in Tissue. 



This is practically the same as the method for cover-glass 

 preparations. 



The section is stained in anilin-water gentian-violet (Koch- 

 Ehrlich) diluted with one-third its volume of water. The 

 section remains in this for about ten minutes at the tem- 

 perature of the incubator. From this it is taken out and 

 washed alternately in Gram's iodine solution and alcohol until 

 all the naked-eye color has been extracted. It is then put 

 into a watery solution .of eosin or Bismarck-brown for one 

 minute, again washed in alcohol a few seconds, and then put 



