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Human monoclonal antibody-based quantification of group 2 grass pollen allergens

Author:
Marth, K  Focke, M  Flicker, S  Valenta, R  


Journal:
JOURNAL OF ALLERGY AND CLINICAL IMMUNOLOGY


Issue Date:
2004


Abstract(summary):

Background: Grasses belong to the most potent allergen sources worldwide. Group 2 grass pollen allergens are recognized by more than 100 million allergic patients. Objective: The aim was to develop an assay for the specific detection and quantification of group 2 grass pollen allergens. Methods: We have isolated a monoclonal human IgE Fab specific for group 2 grass pollen allergens by combinatorial cloning from lymphocytes of a grass pollen-allergic patient. This Fab was converted into a complete human IgG(1) antibody and used together with rPhl p 2 to develop a competitive ELISA for the specific measurement of group 2 allergens. ELISA plate-bound purified recombinant human Phl p 2-specific IgG(1) is incubated with a constant amount of biotinylated rPhl p 2 competing with increasing concentrations of group 2 allergens to be determined. Defined concentrations of purified rPhl p 2 are used to establish a standard curve. The concentration of unlabeled group 2 allergens can thus be deduced from the displacement of biotinylated rPhl p 2, which can be detected with peroxidase-labeled streptavidin. Results: The competition-ELISA measured rPhl p 2 concentrations ranging from 10 ng/mL to 500 ng/mL and allowed to quantify group 2 allergens from 9 different grass families. The results were in good agreement with immunoblot data. Conclusions: The described assay can be used for standardization of diagnostic and therapeutic vaccines as well as for the quantification of group 2 allergens in environmental samples.


Page:
470---474


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