The hsFcγRI-based ADA assay can serve as alternative screening assay or as orthogonal confirmation method for preclinical and clinical immunogenicity testing of IgG therapeutics lacking Fc effector functions.

During development of biotherapeutics, availability of specific assay reagents is usually limited. The possibility to switch from one ligand binding assay technology to another, while using the same reagents, would be desirable. Here, we report on an Alexa647(®)-labeled monoclonal antibody against digoxigenin (mAb-Alexa647(®)) that enables the detection of digoxigenylated analyte-specific ELISA reagents by Gyrolab(™). In an analysis of non-monoclonal antibody (mAb) and mAb drugs, this approach maintained the dynamic range, accuracy and precision of the standard Gyrolab™ approach using analyte-specific Alexa647(®)-labeled Ab. In a rat PK study, results of our approach, standard Gyrolab™ and ELISA were comparable, with difference values within the incurred sample reanalysis acceptance criteria. Therefore, mAb-Alexa647(®) enables an easy switch between ELISA and Gyrolab™, providing an effective way to benefit from both platforms.

Background: The development of immunogenicity assays for clinical drug candidates targeting soluble proteins is challenging when the soluble target might produce either false-positive or false-negative signals in bridging anti-drug antibody screening assays. A generic soluble target removal protocol that uses a pH-dependent depletion was evaluated. Results: An anti-drug antibody bridging assay with a pH-dependent soluble target depletion step was successfully developed. Endogenous target levels of ∼600 nM could be depleted below 8 pM. The assay was highly drug tolerant and met regulatory requirements. Conclusion: A reagent-independent target depletion protocol can be used for immunogenicity testing in the presence of a soluble target. The generic protocol circumvents common depletion or masking protocols.