Production of Antipeptide Antisera

This unit describes methods used for the chemical coupling of synthetic peptides to carrier proteins, required for the preparation of peptide immunogens. The carrier protein described here is keyhole limpet hemocyanin (KLH) because it is the one most commonly used. However, other proteins may be used in place of KLH, including bovine serum albumin (BSA) and ovalbumin. Coupling may be accomplished as described with MBS, which requires a Cys residue in the peptide, or with glutaraldehyde, EDCI, or BDB. Also included are an assay for detecting free sulfhydryl (SH) groups, a means of calculating coupling efficiency, an immunization schedule, an indirect ELISA, and a method for preparing a peptide affinity column. The final methodology described is the multiple antigen (MAP) system.