Rapid and wash-free anti-drug antibody assay enabled by the complementary nanoluciferase biosensor

Abstract

Biotherapeutics have demonstrated remarkable therapeutic efficacy across a wide range of diseases; however, their clinical application is often hindered by the development of anti-drug antibodies (ADAs). Traditionally, ADA monitoring has relied on ligand-binding assays (LBAs) such as enzyme-linked immunosorbent assays (ELISA) or electrochemiluminescence assays (ECLIA). Although effective, these methods require multiple washing and incubation steps, extending data generation time to several hours or even days. In contrast, homogeneous immunoassays offer a promising alternative due to their "mix-and-read" feature, which significantly reduces operational complexity, time, and cost compared to conventional LBAs. In this study, we explored the feasibility of rapid ADA detection using a wash-free homogeneous assay based on the split nanoluciferase (SplitNluc) biosensor system. Through case studies involving diverse therapeutic modalities including antibody-drug conjugates (ADC), Fc-fusion protein, Fc-less multidomain biotherapeutics (MDB), and single-domain antibody (SdAb), we demonstrated that the SplitNluc platform is a versatile and efficient tool for robust and rapid ADA monitoring. This approach not only streamlines the detection process but also holds potential for broader applications in biotherapeutic development and clinical monitoring.