Optimized purification strategies for the elimination of non-specific products in the isolation of GAD65-specific monoclonal autoantibodies.

Autoantibodies against antigens expressed by insulin-producing β cells are circulating in both healthy individuals and patients at risk of developing Type 1 diabetes. Recent studies suggest that another set of antibodies (anti-idiotypic antibodies) exists in this antibody/antigen interacting network to regulate auto-reactive responses. Anti-idiotypic antibodies may block the antigen-binding site of autoantibodies or inhibit autoantibody expression and secretion. The equilibrium between autoantibodies and anti-idiotypic antibodies plays a critical role in mediating or preventing autoimmunity. In order to investigate the molecular mechanisms underlying such a network in autoimmunity and potentially develop neutralizing reagents to prevent or treat Type 1 diabetes, we need to produce autoantibodies and autoantigens with high quality and purity. Herein, using GAD65/anti-GAD65 autoantibodies as a model system, we aimed to establish reliable approaches for the preparation of highly pure autoantibodies suitable for downstream investigation.