Background: Soluble multimeric target proteins can generate a target-mediated false-positive signal in bridging anti-drug antibody (ADA) assays. A high background signal due to target interference was observed in our anti-REGN-Y antibody assay, and two different strategies were evaluated to mitigate this false-positive signal. Results: Multiple anti-target antibodies were tested and found to be ineffective at reducing target interference, so soluble target receptor and co-factor proteins were used in combination to inhibit the target-mediated signal. These competitive blockers synergistically inhibited target interference and increased target tolerance levels, especially when the assay was performed under mild acidic conditions. A separate approach, target immunodepletion using magnetic beads conjugated with an anti-target antibody, was also effective at mitigating the target-mediated signal, also in combination with mild acidic assay pH. Both methods allowed detection of a true ADA signal in monkey and human post-dose serum samples. Conclusion: These methods provide alternative strategies for mitigating target interference when standard anti-target antibodies are ineffective, with the competitive blocker method being recommended, if possible, due to its higher throughput and easier execution.
Keywords: anti-drug antibody; co-factor; mild acidic assay pH; soluble target receptor; target immunodepletion; target interference.