抗原的设计.doc

Antigen Design Antibodies to small peptides have become an essential tool in life science research, with applications including gene product detection and identification, protein processing studies, diagnostic tests, protein localization, active site determination, protein homology studies and protein purification. While it is quite easy to generate anti-peptide antibodies, it is important to carefully consider the ultimate use for the antibody and the sequence used to ensure success. This tech sheet will briefly explore peptide selection and design, coupling strategy, and carrier proteins which are important factors in anti-peptide antisera generation. Serum purification will also be discussed. Peptide Selection and DesignThe first step in the process is the selection of the appropriate peptide sequence. At this step the ultimate use for the antibody must be considered. If the antibody is needed to probe a specific protein domain then the choice is simple. For example, if one is studying proteolytic processing of an N-terminal precursor, antibodies against the N-terminal region of interest would be raised. Likewise if the goal is to monitor the phosphorylation state of a specific sequence, antibodies to the phosphorylated sequence can be used. If the goal is to raise antibodies that will recognize the protein in its native state, the problem becomes more complex. Anti-peptide antibodies will always recognize the peptide. However, the same antibody may not recognize the sequence within the folded intact protein. Sequence epitopes in proteins generally consist of 6-12 amino acids and can be classified as continuous and discontinuous. Continuous epitopes are composed of a contiguous sequence of amino acids in a protein. Anti-peptide antibodies will bind to these types of epitopes in the native protein provided the sequence is not buried in the interior of the protein. Discontinuous epitopes consist of a group of amino acids that are not contiguous but are brought