84 



Scientific Proceedings (hi). 



2. Warthin and Starry's Silver- Agar Cover-glass Method. 



1. Fix tissue in 4 per cent, neutral formol. 



2. Imbed in paraffin (absolute alcohol, xylol, paraffin). 



3. Cut, mount sections on cover-glasses with albumin fixative. 



4. Remove paraffin (xylol, alcohol, water). 



5. Rinse cover-glass with section in 2 per cent, silver nitrate; 

 cover wet section with another perfectly clean cover-glass, so 

 that they are held together by capillary attraction; then place 

 them carefully in a bottle of 2 per cent, silver nitrate, and place 

 in incubator for 30 to 60 minutes; then remove the silver nitrate 

 and separate cover-glasses. They must not separate while in the 

 solution. 



6. Place cover-glass with section in the following reducing 

 mixture : 



2 per cent, silver nitrate solution 3 cc. 



Warm glycerin 5 cc. 



Warm 10 per cent, aqueous gelatin 5 cc. 



Warm 1 Yi per cent, agar suspension 5 cc. 



5 per cent, aqueous hydroquinone solution 2 cc. 



7. Reduce until section is a light reddish brown (several 

 seconds) ; remove and rinse in 5 per cent, sodium hyposulphite 

 solution. 



8. Rinse in distilled water. Clean off with a cloth any pre- 

 cipitate on the cover-glass. 



9. Absolute alcohol-xylol-balsam. 



The mounted section should have a light reddish brown color; 

 if too deep brown or black the spirochetes will not be sufficiently 

 contrasted. They should appear dark reddish brown to jet 

 black against a very light brown background. 



In some cases of poor fixation of the tissue with poor staining 

 of the spirochetes we have found that if the sections before being 

 put into the silver solution are treated for several minutes in a 

 I per cent, solution of uranium or copper nitrate, or a 0.5 per 

 cent, solution of ferric alum, the tissue-reaction is changed in 

 some way so that good staining results. These solutions do not 

 act uniformly in all cases. After their use the section must be 

 thoroughly washed in distilled water before it is put into the 

 silver nitrate solution. 



