title: Epitope mapping by a method that requires no amino acid sequence information.
abstract: A simple, rapid method of epitope mapping has been developed that avoids the often cumbersome requirement of obtaining amino acid sequence information. The protein antigen is digested with various concentrations of carboxypeptidase into a nearly continuous series of polypeptides of different molecular weights, all containing a common N-terminus. The peptides are separated by polyacrylamide gel electrophoresis and then transferred to nitrocellulose paper. After developing the blot with the antibody to be mapped, a nearly continuous stain is observed extending from the position of the intact antigen to the molecular weight of the smallest N-terminal fragment still containing the antibody's epitope. By noting the molecular weight where the stain terminates, the position of the epitope relative to the N-terminus can be determined. Using this methodology, and taking special precautions to inhibit all endoproteinases in the carboxypeptidase preparation, the previously mapped epitopes of six nonoverlapping antibodies to the erythrocyte anion transporter were confirmed.
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D043424:Carboxypeptidase B; D002268:Carboxypeptidases / Q000276:immunology; D043402:Cathepsin A; D000939:Epitopes / Q000032:analysis*; D008970:Molecular Weight; D010179:Pancreas / Q000201:enzymology; D012441:Saccharomyces cerevisiae / Q000201:enzymology; D029701:Saccharomyces cerevisiae Proteins; D013552:Swine
chemical_list: D000939:Epitopes; D029701:Saccharomyces cerevisiae Proteins; D002268:Carboxypeptidases; D043402:Cathepsin A; C499283:PRC1 protein, S cerevisiae; C022633:serine carboxypeptidase; D043424:Carboxypeptidase B