Immunologic analysis of monoclonal and immunoglobulin E antibody epitopes on natural and recombinant Amb a 1.
BACKGROUND
Amb a 1 is the major allergen from ragweed pollen and more than 90% of ragweed-allergic patients react with this protein. Although Amb a 1 was cloned and sequenced in 1991, little is known of the specificity of anti-Amb a 1 antibodies or of the immunologic properties of the recombinant allergen.


OBJECTIVE
To compare binding of monoclonal antibodies (mAb) and IgE antibodies to purified natural Amb a 1 (nAmb a 1) and recombinant Amb a 1 (rAmb a 1).


METHODS
Binding of a panel of anti-Amb a 1 mAb and IgE antibodies to nAmb a 1 or rAmb a 1 was compared by immunoblotting. Chimeric ELISA was used to measure specific IgE to these allergens using 89 ragweed-allergic sera from Austria, Italy, Canada and the United States.


RESULTS
The 8 mAb bound to a 38 kDa Amb a 1 band in ragweed pollen extract and a subset of 5 mAb also bound to the 26 kDa chain of nAmb a 1. A two-site ELISA was developed using a mAb pair, which was approximately 10-fold more sensitive to rAmb a 1. There was a significant correlation between IgE antibody binding to nAmb a 1 and rAmb a 1 (n=89, r=0.79, P<0.001). A subset of approximately 40% of patients showed greater reactivity to nAmb a 1 than to rAmb a 1.


CONCLUSIONS
The data suggest that there is less reactivity of human IgE to rAmb a 1 compared with nAmb a 1. The development of more sensitive, quantitative ELISA for Amb a 1 will require the production of new mAb especially directed against nAmb a 1.
Wopfner
N
N
Christian Doppler Laboratory for Allergy Diagnosis and Therapy, Department of Molecular Biology, University of Salzburg, Austria. nicole.wopfner@sbg.ac.at
Bauer
R
R
Thalhamer
J
J
Ferreira
F
F
Chapman
M
M
D031164:Ambrosia / Q000276:immunology*; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology* / Q000378:metabolism; D015153:Blotting, Western; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000276:immunology*; D006801:Humans; D007073:Immunoglobulin E / Q000276:immunology* / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D010936:Plant Extracts / Q000276:immunology; D010940:Plant Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*; D011058:Pollen / Q000276:immunology; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism
D000911:Antibodies, Monoclonal; D000939:Epitopes; D010936:Plant Extracts; D010940:Plant Proteins; D011994:Recombinant Proteins; D007073:Immunoglobulin E