Microarray immunoassay: Association of clinical history, in vitro IgE function, and heterogeneity of allergenic peanut epitopes(1)
Although antibodies naturally encounter and are selected for their ability to bind the 3-dimensional surface of an antigen, including epitopes that are not made up of sequential amino acids, identification of primary sequences that are recognized by IgE provides important information regarding a protein’s antigenicity.
The majority of sera from peanut-sensitized patients contained IgE to these recombinant proteins. 97% of these patient sera contained specific IgE to at least one of the 3 allergens, whereas 77%, 75%, and 77% recognized Ara h 1, Ara h 2, and Ara h 3, respectively.
The mean signal ratio for Ara h 1 was 0.72 for patient sera versus 0.14 for sera from control subjects not allergic to peanut (P<.0001). For Ara h 2 and Ara h 3, the respective means were 0.77 and 0.86 (P<.0001 for each). Thirty-nine peptides defining 21 major antigenic regions are recognized by more than 5 patients. Ara h 1 contains 13 prominent antigenic regions, whereas Ara h 2 has 4 and N-Ara h 3 also has 4. No single peptide was recognized by more than 35% of the samples tested. However, 67 (87%) of 77 of these samples were positive to at least one peptide. One region of Ara h 1 represented by peptides 73 and 74, corresponding to residues 361 to 385, showed significant IgE binding by using the microarray assay.
The patients with a history of more severe multisystem reactions recognized a significantly greater number of epitopes than those with a history of cutaneous symptoms alone (P<0.01). The same patients have similar recognition of whole recombinant peanut allergens. representative results from one passively sensitized normal nonatopic donor and one atopic donor without peanut allergy. In both cases the high-diversity IgE sera conferred significantly greater sensitivity to peanut extract. Consistent with our hypothesis and the results with passively sensitized human basophils, the more diverse IgE-containing sera conferred greater effector response.
Peanut allergen–specific IgG epitope diversity correlates well with IgE diversity