Epitope Mapping of Region 11–70 of Ovalbumin (<i>Gal </i><i>d</i> I) Using Five Synthetic Peptides

Five successively located peptides, in region 11-70 of the major allergen of ovalbumin (OA) Gal d I (11–19, 20–33, 34–46, 47–55, 56–70), were obtained by manual solid-phase peptide synthesis. These peptides together with the previously reported OA region 1–10 comprise a segment of 70 amino acid residues located at the N-terminal of ovalbumin. The crude peptides were purified by gel filtration and reversed-phase high-performance liquid chromatographies and their sequences were verified. Polyclonal antibodies against the peptides conjugated to carrier protein (BSA) were raised in rabbits. Rocket line immunoelectrophoresis showed that four peptides (20–33, 34–46, 47–55 and 56–70), could deflect OA-line immunoprecipitates. The peptide's affinity to rabbit polyclonal Ig was examined by quantitative precipitation inhibition and the results suggested that an epitope was encompassed in segments 34–55 and 4755. Allergenicity was tested by inhibition of specific IgE binding of ovalbumin, using several sera and a serum pool from 16 egg-allergic patients. The results showed that the allergenicity was distributed over the whole region. These findings suggested that: (a) the region 11–70 of OA seemed not to encompass continuous epitopes; (b) the antigenicity of this region was convincing for peptides 34–46 and 47–55; (c) the allergenicity, though dependent on the patient serum used, was distributed over the whole of region 11–70; (d) peptide 11–19, although weak antigenically was capable of specific IgE inhibition; (e) human and rabbit polyclonal antibodies did not show analogous affinities to the present peptides.