Attachment of C3b to activators of the alternative pathway of complement results in a decrease in regulatory activity expressed by Factor H. Decay-accelerating factor (DAF) and Factor H were found to exhibit quantitatively similar decreases in regulatory activity toward the C3 convertase (C3b,Bb) bound to activators, such as zymosan (Zym) and rabbit erythrocytes (ER), compared to non-activators, such as sheep (ES) and bovine (EB) erythrocytes. Purified DAF and Factor H, in 0.1% NP-40, were assayed by measuring the amount required to release 50% of the radiolabelled Bb in 10 min from C3b,Bb on Zym or cross-linked erythrocytes. The relative effectiveness (i.e. the restriction index, RI) of DAF for accelerating the decay of C3b,Bb on the various particles was: ES (1.0), ER (0.04) and Zym (0.03). The RI for Factor H was: ES (1.0), ER (0.04) and Zym (0.07). The rate of decay of C3b,Bb induced by DAF and Factor H showed similar restriction. The results suggest that the regulatory properties of DAF are reduced if the cells on which it resides become activators of the alternative pathway as a result of transformation, virus infection or surface alteration. These findings may explain reports of dysfunctional DAF on alternative pathway-activating cells.