The specificity and affinity of polyclonal antibodies raised in rabbits against a hapten conjugated to rabbit serum albumin.

In this study, rabbit antisera to hapten-rabbit serum albumin conjugates were investigated regarding antibody titer, affinity, specificity, and affinity distribution. Methyl phosphonic acid p-amino-phenyl 1,2,2-trimethylpropyldiester (MATP) served as model hapten. Four MATP-rabbit serum albumin conjugates with various hapten densities (with and without spacer) were synthesized and used for immunization of rabbits. Antisera were collected over a 130 day-period and characterized with different ELISA methods. We found that immunogens with rabbit serum albumin gave antisera with lower titers, but similar affinity as compared to polyclonal or monoclonal antibodies obtained with non-rabbit protein as carrier protein. Immunogens with a low hapten density led to higher final titers without affecting antibody affinity or specificity. Immunogens containing a bridging group resulted in higher antibody affinity with a changed specificity. The pattern of antibody affinity distribution differed considerably among individual rabbits and showed a non-Gaussian subpopulation distribution.