GENERAL CHARACTERS OF SPIROCILETES. 65 



cover-slip for forty-five to sixty seconds ; wash in dilute 

 solution of sodium carbonate i drop of i per cent, 

 solution in 10 c.c. water). 



Herxheimer and Hubner advise the use of bleu de 

 Nil or bleu de Capri. Modifications of Jenner's or 

 Romanowsky's stain are used by Simonelli and Bandi 

 and by Von dem Borne. The former writers advise the 

 method of May Grunwald, viz., dissolve i grm. of eosin 

 (Grubler) in i litre distilled water, and i gr. of methyl- 

 ene blue (Meister, Lucius and Briining) in a similar 

 quantity of water; mix these solutions, allow to stand 

 for two to seven days, and filter. Dissolve the sedi- 

 ment in pure methyl alcohol, and stain preparations 

 with this solution for four to ten minutes. 



Impregnation with Silver. Bertarelli and Volpino 

 first introduced this mode of staining spirochaetes in 

 tissue, but the modification of their method devised 

 by Levaditi is usually preferred. His original method 

 is as follows: 



Fix fragments of tissue, about i mm. thick, in 10 

 per cent, formol solution; wash in water, and harden 

 in 96 per cent, alcohol ; then place in distilled water till 

 they sink. Next place in the silver solution (silver 

 nitrate 1.5 to 3 per cent.) at a temperature of 33 C. 

 for three to five days. Wash in distilled water, and 

 place in the reducing fluid (pyrogallic acid 2 to 4 per 

 cent., formol 5 c.c., distilled water 100 c.c.) for twenty- 

 four to forty-eight hours. Wash in distilled water, 

 dehydrate, embed and cut (see Plate, Fig. B). A 

 subsequent staining of the sections with Giemsa's stain 

 or with toluydene blue may be carried out. 



A subsequent modification of this method was de- 

 vised by Levaditi and Manouelian, by which the first 

 solution is made of i per cent, silver-nitrate solution 

 with the addition of 10 per cent, pyridine; the tissues 

 5 



