IgE and IgG antibody response to birch pollen antigens were studied by means of immunoblotting experiments testing 58 sera from patients with Type I allergy to birch pollen. 56/58 patients showed IgE antibodies reactive with Bet v I, a 17 kilodalton (kD) pollen protein. 2D-electrophoresis/immunoblot revealed a heterogeneity of that protein. Ten spots (pH 4.9-5.9) could be detected, presumably representing differentially glycosylated isoallergens. In 33/58 patients, there was no evidence of IgE antibodies directed against allergens other than Bet v I. However, in 25/58 of patients' sera, 11 minor allergens (13, 15, 18, 27, 29, 32, 39, 44, 57, and 68 kD) with individual incidences from 1.7% to 17.2% were identified. All proteins were also recognized by the patients' IgG antibodies: in the case of Bet v I recognition was weak, whereas the IgG response to the minor allergens was pronounced. Sera from healthy individuals showed similar IgG antibody responses, but no IgG to the 15, 27, and 29 kD proteins. Our results suggest that IgG directed against minor allergens may function as trapping antibodies in healthy individuals. Too low or lacking amounts of anti-Bet v I IgG may facilitate an allergic reaction.
Two monoclonal antibodies against birch pollen proteins were produced by immunizing BALB/c mice with birch pollen extract. In immunoblotting experiments, antibody BIP 1 reacted with a 17-kilodalton (kD) protein considered to represent the major birch pollen allergen Bet v I. A second monoclonal antibody, BIP 3, reacted with 3 different birch pollen proteins of molecular weights 32, 36 and 68 kD of which the 36- and 68-kD proteins corresponded to minor allergens of birch pollen. Two-dimensional electrophoresis/immunoblotting experiments revealed that BIP 1 reacted with all Bet v I isoallergens, also identified by human IgE antibodies. Using BIP 1 coupled to Sepharose 4B as reverse immunosorbent, Bet v I was obtained in a single-step procedure and characterized as single band by SDS-PAGE.
Sera from 27 birch pollen-allergic patients who had undergone hyposensitization treatment for 22-41 months were studied by immunoblotting before and after therapy, whereby the levels of IgE, IgG and IgG1-4 antibodies directed against the major allergen Bet v I and minor allergens of birch pollen were monitored. The clinical benefit of immunotherapy (IT) was evaluated using a symptom specific questionnaire. In patients with good clinical response (responders, n = 18), as defined by improvement of symptoms, anti-Bet v I IgE antibodies were found to decrease in 10/18 patients (55.5%), whereas in 6/18 (33.3%) no change and in two cases (11.2%) an increase of specific IgE was observed. In the group of patients with unsatisfactory clinical outcome (non-responders, n = 9), 3/9 patients (33.3%) showed a decrease, 3/9 (33.3%) no change and 3/9 (33.3%) an increase in levels of IgE antibodies directed against Bet v I. In the case of minor allergens, 5/18 responders (27.7%) and 8/9 non-responders (88.8%) showed specific IgE before IT. In the responder group, no increase of specific IgE could be observed after IT. In non-responders, however, an increase of IgE directed against minor allergens was seen in 3/9 patients (33.3%). In all patients, regardless of therapeutical success, IT-induced elevated levels of specific IgG, IgG1 and in particular IgG4 directed against Bet v I were found. Regarding minor allergens, a heterogeneous pattern of IgG responses without significant correlation to clinical benefit was observed. Our results indicate that changes in IgG reactivity patterns against Bet v I and minor allergens, as shown by the immunoblot technique, did not correlate with good or bad clinical outcome.
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