ENZYMES OF THE TUBERCLE BACILLUS 



149 



figures being in milligrams of amino acid a nitrogen per 0.5 cc. 

 using asparagin as standard. 



AGE OF AUTOLYSATE 



fT 



One day ^ NHC 



[HC 



fT 



Four days "i NHC 



[HC 



fT 



Eight days .{NHC 



IHC 



T, Test, containing autolysate and elastic tissue. 



NHC, Non-heated control, containing autolysate alone. 



HC, Heated control, containing autolysate (heated) and elastic tissue. 



Experiment II. Connective tissue disintegrating enzyme. This 

 experiment was done exactly as in Experiment I with the excep- 

 tion that the 0.5 gram elastic tissue was replaced by 0.5 gram 

 connective tissue prepared from the capsule of large tubercles. ^ 

 The results of analyses given as the amount of amino acid a 

 nitrogen in milligrams per 0.5 cc. of solution are tabulated in 

 the following table. 



5 In order to prepare the connective tissue for this experiment rabbits were 

 injected subcutaneously in the back with fat free tubercle bacilli of which 0.05 

 gram was used for each site of injection. In order to make a uniform suspension 

 a 10 per cent starch paste was prepared with sterile boiling water and to every 

 5 cc. was added 0.05 gram tubercle bacilli. The bacilli were well mixed with the 

 starch paste by stirring and the entire amount injected subcutaneously while 

 warm by means of a Murphy glycerine syringe or other pressure syringe. After 

 two to three months a large firm tubercle had formed at the site of injection and 

 was removed. After separating all muscle, blood, and caseous material from the 

 thick connective tissue capsule of the tubercle, the latter was well washed with 

 physiological salt solution, then ground up fine in a meat grinder and washed 

 12 times with salt solution or until no more turbidity appeared in the salt solu- 

 tion upon vigorous shaking. The connective tissue was then heated for one- 

 half hour at 90°C. to destroy the enzymes and again washed with salt solution. 

 The connective tissue thus prepared was preserved for use in salt solution with 

 chloroform and toluene. 



