Capacity of purified peanut allergens to induce degranulation in a functional in vitro assay: Ara h 2 and Ara h 6 are the most efficient elicitors
- 9 July 2009
- journal article
- Published by Wiley in Clinical and Experimental Allergy
- Vol. 39 (8), 1277-1285
- https://doi.org/10.1111/j.1365-2222.2009.03294.x
Abstract
Peanut is a most common and potent food allergen. Many peanut allergens have been characterized using, in particular, IgE-binding studies. We optimized an in vitro functional assay to assess the capacity of peanut allergens to degranulate humanized rat basophilic leukaemia cells, RBL SX-38 cells, after sensitization by serum IgE from peanut-allergic patients. We thus compared the activity of the main peanut allergens, i.e. Ara h 1, Ara h 2, Ara h 3 and Ara h 6, purified from roasted peanut. Sera of 12 peanut-allergic patients were collected and total and peanut-specific IgE were measured. They were used to sensitize RBL SX-38 cells and the degranulation was induced by incubation with ranging concentrations of a whole peanut protein extract or of purified peanut allergens. The mediator release was quantified by the determination of beta-hexosaminidase activity in the supernatant. The intensity of the degranulation was expressed as maximum release and as EC50, corresponding to the dose of allergen that induced 50% of the maximum release. For each serum, only 10 IU/mL of human IgE was necessary to sensitize the cells and obtain an optimal degranulation. With all the allergens, the release was positively correlated with the concentration of allergen-specific IgE in the serum used to sensitize the cells. The medians of EC50 obtained for Ara h 2 and Ara h 6 were 2.1 and 2.8 pm, respectively, while they were much higher for Ara h 3 and Ara h 1 (65 and 150 pm, respectively). The RBL SX-38 release assay proved to be sensitive, specific and reproducible. It allowed the comparison of the degranulation potential of different peanut allergens. For all the sera tested, Ara h 2 and Ara h 6 were more potent than Ara h 1 or Ara h 3.Keywords
This publication has 35 references indexed in Scilit:
- Children with peanut allergy recognize predominantly Ara h2 and Ara h6, which remains stable over timeClinical and Experimental Allergy, 2007
- Lounging in a lysosome: the intracellular lifestyle of Coxiella burnetiiCellular Microbiology, 2007
- Regulation of allergy by Fc receptorsCurrent Opinion in Immunology, 2005
- The prevalence of food hypersensitivity in an unselected population of children and adultsPediatric Allergy and Immunology, 2005
- Comparative potency of Ara h 1 and Ara h 2 in immunochemical and functional assays of allergenicityClinical Immunology, 2005
- Development of a functional in vitro assay as a novel tool for the standardization of allergen extracts in the human systemAllergy, 2005
- Purification and immunoglobulin E‐binding properties of peanut allergen Ara h 6: evidence for cross‐reactivity with Ara h 2Clinical and Experimental Allergy, 2005
- Relevance of Ara h1, Ara h2 and Ara h3 in peanut‐allergic patients, as determined by immunoglobulin E Western blotting, basophil–histamine release and intracutaneous testing: Ara h2 is the most important peanut allergenClinical and Experimental Allergy, 2004
- Quantitative determination of total and specific human IgE with the use of monoclonal antibodiesJournal of Allergy and Clinical Immunology, 1986
- Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4Nature, 1970