Monoclonal antibodies against the major allergen of Plantago lanceolata pollen, Pla 1 1: affinity chromatography purification of the allergen and development of an ELISA method for Pla 1 1 measurement

Background: Plantngo lanceolata (English plantain) pollen is a relevant cau se of pollinosis in temperate regions. The major allergen of this pollen, P la 1 1, is recognized by the specific IgE from more than 80% of plantain-se nsitive patients. It displays significant sequence homology with the major olive-pollen allergen Ole e 1. The objective was to develop a monoclonal an tibody-based ELISA to quantify Pla 1 1, and to assess the correlation of Pl a 1 1 content with the biologic activity of plantain pollen extracts. We al so aimed to establish the specificity of the monoclonal antibodies against the potentially cross-reactive allergen Ole e 1, and to investigate the pre sence of Pla 1 1-like proteins in psyllium and melon that have been reporte d to cross-react with P. lanceolata pollen. Methods: After fusion of myeloma cells with spleen cells from a BALB/c mous e, two Pla 1 1-specific monoclonal antibodies secreting hybridomas were sel ected, and the antibodies characterized. One of them (2A10) was used as the capture antibody in an ELISA for Pla 1 1 quantitation. An anti-P. lanceola ta rabbit serum was used as the second antibody. Pla 1 1 was purified by im munoaffinity chromatography and used as the standard in the assay. Results: The ELISA developed was highly reproducible and sensitive, with a detection limit of 0.1 ng/ml, and a practical working range of 0.4-12 ng/ml . The specificity was demonstrated against a large battery of allergens, in cluding Ole e i. The concentration of Pla 1 1 was measured in 19 extracts o f P. lanceolata pollen, and a good correlation was observed between the Pla 1 1 content and the allergenic activity of the extracts. Pla 1 1 was not d etected in psyllium or melon extracts. Conclusions: The results prove the usefulness of the Pla 1 1-ELISA for the standardization of extracts of P. lanceolata pollen intended for clinical u se.