The Glycerin Method 95 



1. Transfer to the stain from water. 



It is seldom necessary to stain longer than 10 minutes. As a 

 rule, it is better to dilute the stain (about Ic.c. tolOc.c. of distilled 

 water) and allow it to act for 10 hours or over night. 



2. Wash in water, 20 minutes. 



3. 10 per cent glycerin until sufficiently concentrated. 



4. Mount and seal. 



Eosin is a good stain for many algae and fungi, when sharp out- 

 lines rather than cell contents are to be brought out. After the 

 material has been fixed and washed in water, stain in an aqueous 

 solution of eosin for 12 to 24 hours. Wash in water until the stain 

 is about right. Since subsequent processes will extract a little 

 more of the stain, the washing in water must stop a little before the 

 desired differentiation has been secured. Place in 1 per cent acetic 

 acid for a few minutes to fix the stain. Then place in 10 per cent 

 glycerin containing about 1 per cent acetic acid, and allow the 

 glycerin to concentrate. The acetic acid is to prevent the stain 

 from washing out. When the glycerin has reached the proper con- 



t 



centration, mount and seal as before. 



The following is a rapid method for forms like Eurotium and 

 Penicillium: Fix in 100 per cent alcohol about 2 minutes; stain 

 in aqueous eosin 5 minutes; wash in water about 1 minute; fix 

 in 1 per cent acetic acid 1 minute; then mount directly in 50 per cent 

 glycerin to w^hich about 1 per cent acetic acid has been added. It is 

 hardly worth while to try this method with forms which have large 

 cells; they are almost sure to collapse. If a form like Eurotium passes 

 through the earlier processes without danger, but collapses when 

 put into the 50 per cent glycerin, put it into the 10 per cent glycerin 

 and allow the glycerin to concentrate. 



Mounting without Fixing or Staining. It is sometimes desirable 

 to retain the natural color of an object. The chlorophyll green can 

 usually be preserved by mounting directly in glycerin without any 

 previous fixing. Other colors also are often preserved in this way. 

 Moss protonema make beautiful preparations by this method. If 

 possible, select protonema showing the very young moss plants. The 

 brown protonema and brown bulbils preserve their color perfectly. 

 W T ash the dirt away from the protonema, which is then placed in 10 



