HEMOLYSINS. 261 



The heating of serum to 56 C. provides one * Exper ._ 

 means of apparent isolation of the amboceptor ment in tiie 

 from the complement, but this is not a true isola- 

 tion in that complement is merely rendered inac- 

 tive by the heat rather than totally eliminated. 



Ehrlich and Morgenroth devised a method by 

 which the amboceptors may be separated from a 

 fresh immune serum without in any way injuring 

 the complement. This is accomplished by per- 

 forming the binding experiment, already alluded 

 to, at a low temperature. The serum, containing 

 both amboceptors and complement, is cooled to 

 C. or slightly above, by means of a freezing mix- 

 ture of salt and ice. A suspension of the homolo- 

 gous corpuscles is cooled to the same point, the 

 serum is added and the mixture maintained at 

 to 4 C. for from fifteen to twenty minutes. At the 

 end of this time the sensitized cells are removed 

 by immediate centrifugation at a low temperature, 

 and are washed entirely free from serum by the 

 use of ice-cold salt solution. If the low tempera- 

 ture has been adhered to rigorously and the work 

 done quickly, the corpuscles are not laked during 

 the manipulations in spite of the presence of both 

 amboceptors and complement. Furthermore, the 

 washed sensitized cells remain intact even when 

 their temperature reaches that of the thermostat, 

 whereas if some fresh normal serum or the serum 

 from which the amboceptors were absorbed is 

 added, they undergo hemolysis as readily as when 

 treated with the active immune serum. The 

 original immune serum is now a solution of com- 

 plement, and fresh corpuscles which are added to 

 it are not dissolved because of the absence of ambo- 

 ceptors. 



