Volume 4 Supplement 2
High density IgE recognition of the major grass pollen allergen, Phl p 1, revealed with single chain IgE antibody fragments obtained by combinatorial cloning
© Madritsch et al; licensee BioMed Central Ltd. 2014
Published: 17 March 2014
Grass pollen is one of the most potent and frequently recognized allergen sources. The timothy grass pollen allergen Phl p 1 belongs to the group 1 of highly cross-reactive grass pollen allergens with a molecular weight of approximately 25-30 kDa. Group 1 allergens are recognized by more than 95% of grass pollen allergic patients. A major mechanism of allergic inflammation is the cross-linking of mast cell and basophil-bound IgE antibodies by allergens.
Here we investigated the IgE recognition of Phl p 1 using allergen specific single chain IgE antibody fragments (IgE-ScFvs) obtained by combinatorial cloning from a combinatorial IgE-ScFv library constructed from PBMC of a grass pollen allergic patient.
Using phage display, two Phl p 1-specific ScFv with high specificity and affinity for Phl p 1 were isolated and their binding sites were found to be localized with synthetic Phl p 1-derived peptides in close vicinity at the N-terminus of the allergen. Moreover, we could show by surface plasmon resonance experiments that both IgE-ScFvs could simultaneously bind to Phl p 1 without notable steric hindrance. Even when we used a combination of the two IgE-ScFvs and an additional human Phl p 1-specific IgE, no relevant inhition of allergic patients polyclonal IgE binding to Phl p 1 could be achieved, indicating high density IgE recognition of the Phl p 1 allergen by multiple IgE antibodies.
Our results indicate that allergic patients IgE antibodies can bind in an unusual density without steric hindrance to Phl p 1 which may explain the high allergenic activity of this allergen.
This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.