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An extract of dry-roasted commercial peanut mix (CPE) was examined for allergenic activity in peanut-sensitive individuals, using skin tests and radioallergosorbent (RAST) assays. Proteins in the extract were characterized by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting. The proteins were electroeluted in three fractions in the ranges 15-25, 26-58 and 65 kDa. The 15-25 kDa molecular weight fraction produced the most reactive skin tests in peanut-sensitive subjects and was chosen for monoclonal antibody production. Six hybridoma cell lines secreting peanutspecific antibodies of the IgM isotype (kappa light chain) were produced. Immunopurified CPE proteins were then subjected to SDS-PAGE, resulting in five major bands with approximate molecular weights of 14, 25, 38, 40 and 44 kDa. Immunoblotting of these separated proteins revealed: (1) three bands with approximate molecular weights of 38, 44 and 65 kDa, which bound IgE from peanut-sensitive patients; and (2) that the monoclonal antibodies recognized epitopes in bands at approximate molecular weights of 12, 14, 23 and 25 kDa. RAST inhibition assays showed that the affinity-purified proteins were able to inhibit the binding of serum IgE from peanut-allergic individuals to solid-phase CPE.

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