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Front Neurol. 2017 Jul 6;8:305. doi: 10.3389/fneur.2017.00305. eCollection 2017.

Development and Validation of an Enzyme-Linked Immunosorbent Assay for the Detection of Binding Anti-Drug Antibodies against Interferon Beta.

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Medical Faculty, Department of Neurology, Heinrich-Heine-University, Duesseldorf, Germany.
Sanofi-Aventis, Deutschland GmbH, Frankfurt am Main, Germany.
Neuroimmunology Laboratory, DMSC, Department of Neurology, Rigshospitalet, Copenhagen, Denmark.
Department of Clinical Neuroscience, Karolinska Institutet, Stockholm, Sweden.
Department of Neurology, Innsbruck Medical University, Innsbruck, Austria.
INSERM 996, University Paris-Sud, Paris, France.
Merck NBE Bioanalytics, Torino, Italy.
Centre for Neuroscience and Trauma, Blizard Institute, Queen Mary, University of London, London, United Kingdom.
Department of Neurology, University Hospital of Cologne, Cologne, Germany.



To 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.


A 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.


The 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.


An 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.


anti-drug antibodies; biotherapy; enzyme-linked immunosorbent assay; interferon beta; multiple sclerosis

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