Multiplex microbead measurements for the characterization of cat and ragweed allergen extracts.

Nicolette C deVore, Susan Huynh, Ekaterina N Dobrovolskaia, Jay E Slater

Center for Biologics Evaluation and Research, US Food and Drug Administration, Bethesda, Maryland, USA.

Annals of allergy, asthma & immunology : official publication of the American College of Allergy, Asthma, & Immunology 2010 Nov

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Current assays for allergen extracts can measure either overall potency or the levels of individual allergens. To develop a multiplex allergen extract potency assay (MAEPA) for allergen extracts that can concurrently measure individual allergens and characterize the overall allergen levels in the mixture. Six anti-Fel d 1 and 6 anti-Amb a 1 recombinant antibodies were generated and were covalently bound to carboxy-labeled beads. Antibody-bound beads were then used to measure Fel d 1 and Amb a 1 levels in commercial cat hair and short ragweed pollen (SRP) extracts, respectively, using bead-based flow cytometry. These major allergen levels were compared with those obtained using a conventional antibody-based method. Allergen levels were calculated by comparing the half-maximal effective concentrations of dose-response curves analyzed using 4-parameter fits. Bead-antibody pairs were tested to determine whether the presence of additional bead-antibody pairs affected the apparent potency of the extract. Allergen contents of cat hair and SRP extracts determined using the MAEPA and anti-Fel d 1 and anti-Amb a 1 antibodies were comparable with potencies determined using conventional methods. Cross-interference from the concurrent use of multiple beads was minimal. Six lots of cat hair extract and 6 lots of SRP extract were tested. The MAEPA, a bead-based assay using recombinant antibodies, accurately determined Fel d 1 levels in cat hair allergenic extracts and Amb a 1 levels in SRP extracts. The results of this assay are reproducible and are consistent with data obtained using conventional methods. Copyright © 2010 American College of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

Citation

Nicolette C deVore, Susan Huynh, Ekaterina N Dobrovolskaia, Jay E Slater. Multiplex microbead measurements for the characterization of cat and ragweed allergen extracts. Annals of allergy, asthma & immunology : official publication of the American College of Allergy, Asthma, & Immunology. 2010 Nov;105(5):351-8