In order to understand enzyme P-450 mediated metabolism of endobiotics and xenobiotics we are characterizing the antigenic sites (epitopes) of cytochrome P450. Each monoclonal antibody (MAb) is a homogeneous reagent that is highly specific for a single epitope. We are characterizing cross- reactivity of MAbs to different forms of cytochrome P450 with recombinantly expressed P450s of different species and preparing MAbs against purified rat cytochrome P450 1A2. The existing library of mouse MAbs raised against 3-methylcholanthrene and phenobarbital-induced rat liver cytochrome P450 was tested for cross- reactivity to other species. Enzymatically active cytochrome P450 (mouse 1A1, 1A2, rat 2B1, and human 1A2 and 2B1) were expressed in TK- and Hep G2 human cell lines using the vaccinia virus expression system. Preliminary results of cross-reactivity studies inside subfamilies 1A1/2 and 2B1/2 suggest that MAbs raised against rat P450 react with recombinantly expressed rat and mouse P450, but not with human. In order to extend our library of MAbs we are preparing mouse IgG to purified rat cytochrome P450 1A2. One clone was obtained which was found to react specifically with rat P450 1A2.
