Abstract
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 |
|---|---|
| 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 |
Keywords
- Animals
- Antibodies
- Antibody Specificity
- Chromatography, Affinity
- Electrophoresis, Polyacrylamide Gel
- Enzyme-Linked Immunosorbent Assay
- HLA-DQ Antigens
- Immunoblotting
- Peptides
- Rabbits
- Rats
- Receptors, Somatotropin