Isolation 777 



transferring them to a solution of sulphuric and oxalic 

 acids 



Concentrated sulphuric acid 2 drops 



Five per cent, oxalic acid solution i drop 



Distilled water 10 c.c. 



for five seconds, then to absolute alcohol, xylol, etc. The 

 bacilli appear dark blue upon a paler ground. This method 

 gives very good results, but has been largely superseded by 

 the use of Kiihne's carbol-methylene-blue : 



Methylene-blue i .5 



Alcohol 10.0 



Five per cent, aqueous phenol solution 100.0 



Kiihne stains the section for about half an hour, washes it 

 in water, decolorizes it carefully in hydrochloric acid (10 

 drops to 500 c.c. of water), immerses it at once in a solution 

 of lithium carbonate (8 drops of a saturated solution of lithium 

 carbonate in 10 c.c. of water), places it in a bath of distilled 

 water for a few minutes, dips it into absolute alcohol col- 

 ored with a little methylene-blue, dehydrates it in anilin 

 oil containing a little methylene-blue in solution, washes 

 it in pure anilin oil, not colored, then in a light ethereal oil, 

 clears it in xylol, and finally mounts it in balsam. 



Vital Resistance. The organism grows only between 

 25 and 42 C. It is killed by exposure to 60 C. for two 

 hours, or to 75 C. for one hour. Sunlight kills it after 

 twenty-four hours' exposure. Thorough drying destroys it in 

 a short time. When planted upon culture-media, sealed, and 

 kept cool and in the dark, it may be kept alive for months 

 and even years. Exposure to i per cent, carbolic acid de- 

 stroys it in about half an hour; i : 1000 bichlorid of mercury 

 solution, in about fifteen minutes. According to Hiss and 

 Zinsser, it may remain alive in the water of horse-troughs 

 for seventy days. 



Isolation. Attempts at the isolation of the glanders 

 bacillus from infectious discharges by the usual plate method 

 are apt to fail, on account of the presence of other more 

 rapidly growing organisms. 



The best method of isolation seems to be by infecting an 

 animal and recovering the bacillus from its tissues. 



The guinea-pig, being a highly susceptible as well as a 

 readily procurable animal, is appropriate for the detection 



