Identification of potential allergens in white oak (Quercus alba) pollen by immunoblotting

Aqueous extracts of white oak pollen were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and transferred onto nitrocellulose membranes. The nitrocellulose membranes were blocked with phosphate-buffered saline 15% nonfat dry milk, incubated with dilutions of sera from atopic or control subjects, and probed with a radiolabeled or peroxidase-labeled antihuman IgE. The IgE binding bands were detected by autoradiography or enzymatic reaction; 45 to 50 protein bands were observed in silver-stained gels. IgE from 30 of the 38 sera tested from oak-sensitive subjects bound to 23 bands with molecular weights (MWs) between 106 to 108 kd (band 1) and 13.2 to 15.2 kd (band 23). No band was recognized by sera of every patient. Band 5 (MW 74.0 to 77.9 kd) and band 21 (MW 16.2 to 17.7 kd) were recognized by 71% of the patients' sera. Multiple bands were recognized by 30% to 50% of the sera tested. All patients who were skin test positive to oak by prick testing had positive immunoblots. Of 12 patients positive by intradermal skin testing, only four patients had positive immunoblots. The average number of allergens recognized by a single patient was 6.6. The maximum number of allergens to which any individual reacted was 18; the minimum number was one. Extracts separated under nonreducing conditions resulted in aggregates that did not enter the polyacrylamide gel. Of the protein that did enter the gel, the higher MW species elicited banding patterns similar to patterns observed under reducing conditions, whereas lower MW IgE binding bands were lost. These data suggest that the extractable proteins of white oak pollen contain multiple proteins that are potentially allergenic.