184 



INFECTION AND RESISTANCE 



fractions, and the writer has noticed similar failures but has always 

 been able to obtain eventual separation by sufficient prolongation of 

 the dialysis. Because of the occasional difficulties and because of 

 the time-consuming and inconvenient nature of the method other 

 means of separation have been devised. The one used with success 

 by many workers has been that introduced by Sachs and Altmann, 64 

 namely, precipitation of the sera with weak hydrochloric acid -g^ 

 to -j^. Liefmann has separated the components by precipitation of 

 the globulins by the introduction of CO 2 . In carrying out this 

 method, Fraenkel 65 has found it advantageous to dilute the serum 

 ten times with distilled water, then allowing the CO 2 to flow in at 

 low temperatures. It is likely that any of the usual methods of 

 globulin separation will serve for complement partition. The salt- 

 ing out methods are, however, extremely inconvenient because of the 

 prolonged dialysis subsequently necessary to remove the salts. 



The inactivation of complement or alexin by the addition of 

 salts or by splitting is, very apparently, a temporary inactivation in 

 which prompt restitution can be practiced by bringing back original 

 conditions either by dilution to isotonicity or by reconstruction of 

 the divided substance, respectively. Heating to 56 C., the simplest 

 and most commonly employed method of inactivations was, until of 

 late, regarded as an irreversible process, the complement being irre- 

 trievably destroyed in the procedure. Gramenitski 66 has recently 

 carried out experiments which seem to show that this opinion is 

 erroneous. His experiments were suggested by the fact, observed by 

 Bach and Chodat, 67 that certain oxydases and diastases may spon- 

 taneously regain some of their activity after inactivation by heat. 

 His work with complement indicated a similar gradual return to an 

 active condition after moderate heating. The great theoretical im- 

 portance of this observation will justify our insertion of one of 

 Gramenitski's protocols. 



Experiment 1. Complement 10 times diluted was heated to 56 

 C. for 7 minutes. It was then tested against sensitized beef blood 

 at varying intervals as follows: 



64 Sachs and Altmann. Cited from Sachs in "Kolle u. Wassermann Hand- 

 buch," Vol. 2, p. 877. 



65 Fraenkel. Zeitschr. /. 7mm., I, Vol. 8, 1911. 



66 Gramenitski. Biochem. Zeits., Vol. 38, 1912. 



67 Bach and Chodat. Cited from Gramenitski, loc. cit., p. 511. 



