Establishment of a reporter gene-based assay to determine ADCC activity of anti-tetanus toxin monoclonal antibody.

Tetanus remains a serious health concern, and therapeutic monoclonal antibodies targeting the tetanus toxin C-fragment (TT-C) require reliable methods to assess antibody-dependent cell-mediated cytotoxicity (ADCC) activity. In this study, we describe the development of a validated luciferase reporter ADCC assay using a stable Flp-In CHO cell line stably expressing high levels of TT-C as target cells. The generated CHO cell line exhibited stable and high-level TT-C expression, with 99.36% TT-C-positive cells. We found the assay produced a highly reproducible sigmoidal dose-response curve (R2 > 0.99), and method validation demonstrated high specificity, precision (intermediate precision RSD = 13.90%), and accuracy (spike recoveries between 99.99% and 103.18%). This robust assay provides a reliable platform for functional characterization, potency evaluation, and quality control of anti-tetanus toxin neutralizing monoclonal antibody.