STUDIES ON IMMUNE HEMOLYSIS: PREPARATION OF A STABLE AND HIGHLY REACTIVE COMPLEX OF.SENSITIZED ERYTHRO- CYTES AND THE FIRST COMPONENT OF COMPLEMENT (EAC'I); INACTIVATION OF CELL-FIXED C'l BY SOME COMPLEMENT REAGENTS* BY PAUL G. KLEIN, M.D

The interaction of sensitized sheep erythrocytes (EA) with complement (C') leads to hemolysis of the red cells in a sequence of several reaction steps. Levine and Mayer (1) have established that the components C'I and C'4 first interact with the sensitized cells to form an intermediate complex EAC'1,4. Whether the transfer of hemolyticaUy active C'1 to sensitized erythrocytes is dependent upon previous or simultaneous fixation of C'4, or whether the fixation of C'1 precedes and then mediates the subsequent action of C'4 has not been satisfactorily determined. Pillemer and coworkers (2) were unable to transfer hemolytically active C'1 to sensitized cells in the absence of C'4. They found, after interacting hydrazine-treated complement with sensitized cells, that not only was the supernatant serum devoid of C'1 activity, but also that there was no detectable C'1 activity on the resultant cells. Becker and Wirtz (3), on the other hand, were able to obtain a reactive EAC'I by exposing keavily sensitized erythrocytes to hydrazine-treated guinea pig serum. Laporte et al. (4) exposed an EA preparation to the euglobulin fraction (midpiece) of hydrazine-treated guinea pig serum (R4), and found that the cells were converted into EAC'I. The experiments reported by the latter two groups of investigators indicate that it is indeed possible to produce an EAC'I in the absence of C'4 but only under certain empirically determined circumstances, the mechanisms of which are poorly understood. It appears that the interaction of hydrazine-treated guinea pig serum with sensitized erythrocytes results in deterioration of the complex EAC'I as it is being formed. This capacity of hydmzlne-treated guinea pig serum apparently can either be overcome by using heavily sensitized erythrocytes or eliminated by splitting the hemolytlcally active C~1 into the midpiece fraction of the R4.