A novel affinity purification method to isolate peptide specific antibodies
Research output: Contribution to journal › Journal article › Research › peer-review
Site-specific, high affinity polyclonal antisera are effectively and successfully produced by immunizing rabbits with synthetic peptides. The use of these antisera in subsequent immune analysis is often limited because of non-specific binding. We describe a new and simple method to effectively affinity-purify anti-peptide antibodies. To test our system, rabbits were immunized with model peptides representing sequences of the putative rabbit growth hormone receptor and several HLA-DQ beta-chain molecules. Polystyrene plastic beads were coated with peptides. Immune serum was incubated with the beads and after a wash step the bound antibodies were eluted in 1 M acetic acid. The eluted material was composed predominantly of intact immunoglobulin as evidenced by the presence of heavy and light chain bands in SDS-PAGE. The eluted antibodies were peptide specific in ELISA and bound only to intact, antigenic protein in immunoblot analyses. The sequence-specific nature of the eluted antibodies was confirmed since binding to the antigenic proteins could be displaced by the immunizing but not by unrelated peptides.
Original language | English |
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Journal | Journal of Immunological Methods |
Volume | 128 |
Issue number | 2 |
Pages (from-to) | 151-7 |
Number of pages | 7 |
ISSN | 0022-1759 |
Publication status | Published - 17 Apr 1990 |
- Animals, Antibodies, Antibody Specificity, Chromatography, Affinity, Electrophoresis, Polyacrylamide Gel, Enzyme-Linked Immunosorbent Assay, HLA-DQ Antigens, Immunoblotting, Peptides, Rabbits, Rats, Receptors, Somatotropin
Research areas
ID: 45574812