STAINING OF SPORES. 135 



about five minutes after the last boiling, the preparation 

 is transferred, without washing in water, into a second 

 watch-crystal containing the following decolorizing solu- 

 tion : 



Absolute alcohol 100 c.c. 



Hydrochloric acid 3 c.c. 



In this solution it is placed, bacteria up, and the vessel 

 is tilted from side to side for about one minute. It is 

 then removed, washed in water, and stained with the 

 methylene-blue solution. The spores will be stained 

 red and the body of the cells will be blue. 



MOELLER'S METHOD FOR STAINING SPORES. A 

 method that has recently been published by Moeller is 

 designed to favor the penetration of the coloring material 

 through the spore membrane by macerating the spores 

 in a solution of chromic acid before staining them. It 

 is as follows : 



The cover-slips are prepared in the usual way, or the 

 fixing may be accomplished with absolute alcohol instead 

 of high temperatures. The preparation is then held for 

 two minutes in chloroform, then washed off in water, 

 then placed for from one-half to two minutes in a 5 per 

 cent, solution of chromic acid ; again washed off in 

 water, and now stained in carbolic fuchsin. In the 

 process of staining, the slip is taken by the corner with 

 the forceps and carbolic fuchsin is dropped upon the side 

 containing the spores. It is then held over the flame 

 until it boils, and then held some distance above the 

 flame for one minute. The staining fluid is then poured 

 off and the preparation is completely decolorized in 5 

 per cent, sulphuric acid, again washed off in water, and 

 finally stained for thirty seconds in the watery methy- 



