Comparative studies on tree pollen allergens. XIV. Characterization of the birch (Betula verrucosa) and hazel (Corylus avellana) pollen extracts by horizontal 2-D SDS-PAGE combined with electrophoretic transfer and IgE immunoautoradiography

The crude aqueous extracts of birch (Betula verrucosa) and hazel (Corylus avellana) pollen were characterized by horizontal isoelectric focusing (IEF) and one-dimensional and two-dimensional sodium dodecyl sulfate polyacrylamide gel electrophoresis (1-D and 2-D SDS/PAGE). Isoelectric focusing reference patterns obtained in pH 3.5 to 9.5 gradient gels contained 31 and 41 bands while the 2-D patterns consisted of 51 and 72 protein spots, respectively. Electrophoretic transfer to nitrocellulose membranes and subsequent 125I-immunoautoradiography demonstrated IgE binding in the region between pH 4.8 and 5.4 for both extracts. The main allergenic reactivities of this pH region were distributed by SDS/PAGE to the molecular weights of 29.5, 17, and 12.5 kilodaltons for birch and 15.5 and 12 kilodaltons for hazel. The high resolving power and the independence of rabbit antibodies were considered the major advantages of the described 2-D methodology as compared with conventional crossed immunoelectrophoretic and radioimmunoelectrophoretic techniques.