Pei Min, Wang Yao, Tang Lei, Wu Weitao, Wang Chunhe, Chen Yi-Li
Department of Antibody Discovery, Shanghai Mabstone Biotechnology, Ltd, Shanghai, China.
Department of Research and Development Center, Dartsbio Pharmaceuticals Ltd, Zhongshan, Guangdong, China.
Bio Protoc. 2022 Oct 5;12(19). doi: 10.21769/BioProtoc.4522.
Bispecific antibodies (BsAbs) are typically monoclonal antibody (mAb)-derived molecular entities engineered to bind to two distinct targets, including two antigens or two epitopes on the same antigen. When compared to parental monoclonal antibodies or combinational therapies, the generated BsAbs have the ability to bridge the two targets and thus may offer additional clinical benefits. Characterizing BsAbs' ability to bind to both targets simultaneously is critical for their biotherapeutic development. A range of bi-functional quantitative bridging assays to enable target-specific capture and detection of binding properties include enzyme-linked immunosorbent assay (ELISA), surface plasmon resonance (SPR), and cell-based flow cytometry. Developing suitable and robust cell-based bioassays is more challenging than non-cell-based binding assays because cell-based assays with complex matrices can be inherently variable and often lack precision. Compared to SPR, ELISA has a rapid setup and readily available method, being widely and extensively applied in almost every laboratory. Here, we describe a dual-target bridging ELISA assay that characterizes the ability of a HER2(human epidermal growth factor receptor 2)/PD-L1(programmed cell death ligand 1) BsAb in binding to both HER2 and PD-L1 simultaneously, a prerequisite for its envisioned mode of action. Graphical abstract.
双特异性抗体（BsAbs）通常是源自单克隆抗体（mAb）的分子实体，经过工程改造以结合两个不同的靶点，包括两种抗原或同一抗原上的两个表位。与亲本单克隆抗体或联合疗法相比，生成的双特异性抗体有能力连接两个靶点，因此可能带来额外的临床益处。表征双特异性抗体同时结合两个靶点的能力对其生物治疗开发至关重要。一系列用于实现靶点特异性捕获和结合特性检测的双功能定量桥接分析方法包括酶联免疫吸附测定（ELISA）、表面等离子体共振（SPR）和基于细胞的流式细胞术。开发合适且稳健的基于细胞的生物分析方法比基于非细胞的结合分析方法更具挑战性，因为具有复杂基质的基于细胞的分析方法本质上可能存在差异且往往缺乏精确性。与SPR相比，ELISA设置快速且方法易于获得，几乎在每个实验室都得到广泛应用。在此，我们描述了一种双靶点桥接ELISA分析方法，该方法表征了HER2（人表皮生长因子受体2）/PD-L1（程序性细胞死亡配体1）双特异性抗体同时结合HER2和PD-L1的能力，这是其预期作用模式的一个先决条件。图形摘要。
