ISOLATION OF THE COLON BACILLUS. 71 



thirds of the closed arm and about one-third of this should 

 be absorbed. The nitrate tube is tested for nitrites by 

 adding a drop of the following solutions in succession: 



A. Sulphanilic acid 5 gram 



Acetic acid (25% sol.) 150.0 c.c. 



B. Naphthylamine chloride .1 gram 



Distilled water 20.0 c.c. 



Acetic acid (25% sol.) 150.0 c.c. 



A red or violet coloration indicates the presence of 

 nitrites. The milk tube is merely heated to boiling over 

 the free flame ; if coagulation occurs the test is considered 

 positive. 



After seventy- two hours incubation at 37 the peptone 

 solution is examined for indol by adding i c.c. of a .02 per 

 cent solution of sodium or potassium nitrite and i c.c. of a 

 i to i solution of sulphuric acid. Both the tube and 

 the reagents should be cooled on ice before mixing, and 

 the tube should be left in a cool place for an hour after- 

 ward to allow time for the characteristic rose- red color 

 of nitroso-indol to develop. The gelatin tube is kept at 

 20 for seven days according to the procedure adopted 

 at the Massachusetts Institute of Technology. It seems 

 undesirable in practice to prolong the test much beyond 

 this point, although some slowly liquefying organisms 

 are doubtless included, which would be thrown out by a 

 longer incubation. The extent of this source of error 

 as well as the relative importance of the various other 

 diagnostic tests is well shown in the following table of 



