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The preparation of bacterial intermediates bearing complement components at various steps of the complement sequence was investigated by suspending immunoglobulin M-opsonized Escherichia coli O111:B4 cells in complement-deficient sera at different temperatures and ionic strengths. The optimal conditions for the formation of the intermediates at Tmax were found to be an ionic strength of 0.091 mu and a temperature of 37 degrees C, except for BAC142, which could be formed equally well at room temperature. In contrast to all the other intermediates, which, once formed at Tmax, were stable in the presence of the whole serum, BAC142 decayed with a half-life of 10 min due to the lability of bound C2. Washing with a buffer of either 0.091 or 0.046 mu did not affect the bacterial intermediates, with the exception of BAC1-3 formed either in the presence of C5-deficient serum or with purified C3 added to BAC142. All the intermediates were found to be stable after incubation in 0.091-mu buffer for 30 min at 37 degrees C.

Kinetics of assembly and decay of complement components on Escherichia coli O111:B4 preparation of stable intermediates