Staining 789 



approximately 0.5 per cent. French eosin, and this is added gradually 

 while the mixture is being stirred until a filtered sample shows the filtrate 

 to be of a pale bluish color with a slight fluorescence. The mixture 

 is allowed to stand for one day and then filtered. The precipitate 

 which has separated is collected on a double piece of filter-paper and 

 dried at room temperature (heating spoils it). When completely 

 dried it can easily be removed from the paper and may then be dissolved 

 without further washing in commercial (not pure) wood alcohol. The 

 solution should be allowed to stand a day, then filtered. The strength 

 of this alcoholic solution is approximately i per cent. To use the stain, 

 one drops upon an unfixed spread enough dye to cover it, permits it 

 to act for three or four seconds, and then pours it off and introduces the 

 glass slowly, spread side down, into clean water, where it is held for 

 another four or five seconds, after which it is shaken to and fro in the 

 water to wash it. It is next dried and examined at once or after mount- 

 ing in balsam. The spirochaetes appear violet in color. 



Ghoreyeb* recommends the following rapid method of 

 staining the organism in smears. A thin spread is to be 

 preferred. No heat fixation is necessary: 



1. Cover the smear with a i per cent, aqueous solution of osmic acid, 

 and permit it to act for thirty seconds. This solution acts as a fixative 

 and mordant. 



2. Wash thoroughly in running water. 



3. Cover the smear with a i : 100 dilution of Liquor plumbi subace- 

 tatis (freshly prepared). Permit it to act for ten seconds. The lead 

 unites with the albumin to form lead albuminate which is insoluble in 

 water. 



4. Wash thoroughly in running water. 



5. Cover the smear with a 10 per cent, aqueous solution of sodium 

 sulphid. This is to act ten seconds, during which the salt transforms 

 the lead albuminate into lead sulphid and causes the preparation to turn 

 brown. The osmic acid when reapplied causes it to become black. 



6. Wash thoroughly in running water. 



The whole process is to be repeated in exactly the same man- 

 ner three times, the washings all being very thorough. The 

 preparation is then dried and mounted in Canada balsam. 

 The micro-organisms and cellular detritus are stained black. 



When serum from a primary sore or other syphilitic lesion 

 is treated by these methods, a number of the spirochaeta ap- 

 pear well stained and a number very palely stained, so that 

 one is in doubt whether there may be many others un- 

 stained, and this seems to be the case, for when similar 

 smears are treated by other methods many more can be 

 found. 



The method of silver incrustation was first employed for 

 the demonstration of the organism in tissues, but Sternt 



" Jour. Amer. Med. Assoc.," May 7, 1910, liv., No. 19, p. 1498. 

 t "Berliner klin. Wochenschrift," 1907, No. 14. 



