POLYCLONAL ANTIBODY CATALYTIC VARIABILITY

We have performed a systematic variability study of polyclonal antibod y catalysis by using five rabbits immunized with the same hapten. Impo rtant results from this work are the following. (1) Similarities were observed in the catalytic polyclonal antibodies derived from all five rabbits. Four of the five rabbits produced polyclonal samples that wer e nearly the same in terms of catalytic activity, whereas the fifth ra bbit, designated as rabbit 2, displayed a somewhat higher level of cat alytic activity. The catalytic activities (as k(cat)/k(uncat)) of thes e polyclonal samples were similar to that from the best murine monoclo nal antibody that had been previously elicited by the same hapten. (2) Titre was not an accurate indicator of polyclonal antibody catalytic activity. (3) A mathematical analysis to describe a distribution of Mi chaelis-Menten catalysts was performed to help interpret our results. (4) Kinetic analysis indicated that the binding parameters of the diff erent samples were remarkably homogeneous, because one or two componen ts were all that were required to fit the on-rate and off-rate data sa tisfactorily. Interestingly, the most active catalytic polyclonal samp le, that from rabbit 2, displayed the slowest off-rate (so slow it cou ld not be measured) and thus the highest overall affinity. (5) Catalyt ic analysis of eluted fractions of antibody from a substrate column in dicated that each polyclonal sample was also relatively homogeneous in terms of catalytic parameters. The main conclusion of our study is th at for this hapten-animal system, the overall catalytic immune respons e is relatively consistent at two levels. Consistent catalytic activit y was observed between the polyclonal samples elicited in the differen t animals, and the elicited hapten-specific polyclonal antibodies were relatively homogeneous in terms of binding and catalytic parameters w ithin each immunized animal. The observed similarities of the catalyti c activity in the different animals is surprising, because the immune response is based on specific binding of antibodies to hapten. There i s no known selective pressure to maintain consistent levels of catalyt ic activity. Our results can therefore be interpreted as providing evi dence that for this hapten there is a fixed relationship between hapte n structure and catalytic activity and/or consistent genetic factors t hat dominate the catalytic immune response.