ArticleAllergy2025
Immunological In Vitro Assay for Quantification of Adjuvanted Allergoids.
Article in Allergy, 2025. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Not yet cited in PubMed.
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6 authors.
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Abstract
backgroundMany IgE-mediated allergic disorders can be treated with allergen immunotherapy (AIT). In order to improve safety and efficacy, some AIT products contain allergen extracts which are chemically cross-linked to generate allergoids and are adsorbed to aluminum hydroxide adjuvant. The modification and adsorption impair accessibility of the protein and quantification of the allergoid content.
methodsAn ELISA-like assay to quantify the allergoid content in adjuvanted grass pollen allergoid AIT products (from here on called: AIT drug products; AIT-DPs) was developed using a fluorescence detection system. The high density of the aluminum hydroxide particles enabled pelleting the antigen complexes by centrifugation. Rabbit anti-grass pollen allergoid sera or a mouse anti-Phl p 5 monoclonal antibody (mAb) was used as the primary antibody. Protein content of the samples was quantified by nitrogen analysis.
resultsHigh specificity of the primary antibodies was confirmed by isoelectric focusing, gel-electrophoresis, and immunoblotting. Performance of the allergoid content assay was demonstrated in grass pollen AIT-DPs with high specificity and low/absent cross-reactivity with tree pollen or mite AIT-DPs. It was used to confirm batch-to-batch consistency and allergoid content of distinct grass pollen AIT-DPs. Overall, in relation to their total protein content, the allergoid content ranged between 0.8 and 2.1 relative to an in-house reference for all grass pollen AIT-DPs, whereas use of mAb revealed product-specific differences in the Phl p 5 amount. Additionally, the assay detects product alteration by heat stress.
conclusionThe described assay is suitable to quantify the allergoid content and quality of allergoids in complex with aluminum hydroxide. It is suitable for animal-free final product testing in vitro, for example, for batch release to ensure the quality of AIT-DPs.
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