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A novel affinity purification method to isolate peptide specific antibodies

Publikation: Bidrag til tidsskriftTidsskriftartikelForskningfagfællebedømt

  • Alan E Karlsen
  • A Lernmark
  • Hans Kofod
  • T Dyrberg
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.
OriginalsprogEngelsk
TidsskriftJournal of Immunological Methods
Vol/bind128
Udgave nummer2
Sider (fra-til)151-7
Antal sider7
ISSN0022-1759
StatusUdgivet - 17 apr. 1990

ID: 45574812