1. Development and Validation of an Enzyme-Linked Immunosorbent Assay for the Detection of Binding Anti-Drug Antibodies against Interferon Beta
- Author
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Kathleen Ingenhoven, Daniel Kramer, Poul Erik Jensen, Christina Hermanrud, Malin Ryner, Florian Deisenhammer, Marc Pallardy, Til Menge, Hans-Peter Hartung, Bernd C. Kieseier, Elisa Bertotti, Paul Creeke, Anna Fogdell-Hahn, and Clemens Warnke
- Subjects
multiple sclerosis ,anti-drug antibodies ,interferon beta ,enzyme-linked immunosorbent assay ,biotherapy ,Neurology. Diseases of the nervous system ,RC346-429 - Abstract
ObjectiveTo develop and validate a method for the detection of binding anti-drug antibodies (ADAs) against interferon beta (IFN-β) in human serum as part of a European initiative (ABIRISK) aimed at the prediction and analysis of clinical relevance of anti-biopharmaceutical immunization to minimize the risk.MethodA two-tiered bridging enzyme-linked immunosorbent assay (ELISA) format was selected and validated according to current recommendations. Screening assay: ADA in serum samples form complexes with immobilized IFN-β and biotinylated IFN-β, which are then detected using HRP labeled Streptavidin and TMB substrate. Confirmation assay: Screen “putative positive” samples are tested in the presence of excess drug (preincubation of sera with 0.3 µg/mL of soluble IFN-β) and percentage of inhibition is calculated.ResultsThe assay is precise, and the sensitivity of the assay was confirmed to be 26 ng/mL using commercially available polyclonal rabbit antihuman IFN-β in human sera as the positive control.ConclusionAn ultrasensitive ELISA for IFN-β-binding ADA testing has been validated. This will form the basis to assess anti-biopharmaceutical immunization toward IFN-β with regards to its clinical relevance and may allow for the development of predictive tools, key aims within the ABIRISK consortium.
- Published
- 2017
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