Targeting Antipeptide Antibodies Toward Cytochrome P450 Enzymes

An approach to raising antibodies is described that can be applied to the majority of cytochrome P450 (P450) enzymes. Its application is limited only by the availability of suitable protein sequence information. The method is based on immunizing animals with synthetic peptides that mimic small regions of target proteins. In practice, this has proved to be a relatively simple, rapid, and effective method of producing antibodies. The antibodies are particularly suited to immunoblotting (Western blotting) and immunocytochemistry. Unlike antibodies produced by other techniques, the epitopes of these antibodies are predetermined, allowing them to be directed toward specific regions of P450. Detailed practical information is described on the selection of regions to target, the synthesis and conjugation of peptides to carrier protein, immunization, and the assessment of the resultant antisera. The approach is illustrated using examples of antibodies targeted against rat and human P450.