State Key Laboratory of Robotics, Shenyang Institute of Automation, Chinese Academy of Sciences, Shenyang 110016, China.
Biochem Biophys Res Commun. 2013 Sep 6;438(4):709-14. doi: 10.1016/j.bbrc.2013.07.114. Epub 2013 Aug 2.
Fc gamma receptors (FcγR), widely expressed on effector cells (e.g., NK cells, macrophages), play an important role in clinical cancer immunotherapy. The binding of FcγRs to the Fc portions of antibodies that are attached to the target cells can activate the antibody-dependent cell-mediated cytotoxicity (ADCC) killing mechanism which leads to the lysis of target cells. In this work, we used atomic force microscopy (AFM) to observe the cellular ultra-structures and measure the biophysical properties (affinity and distribution) of FcγRs on single macrophages in aqueous environments. AFM imaging was used to obtain the topographies of macrophages, revealing the nanoscale cellular fine structures. For molecular interaction recognition, antibody molecules were attached onto AFM tips via a heterobifunctional polyethylene glycol (PEG) crosslinker. With AFM single-molecule force spectroscopy, the binding affinities of FcγRs were quantitatively measured on single macrophages. Adhesion force mapping method was used to localize the FcγRs, revealing the nanoscale distribution of FcγRs on local areas of macrophages. The experimental results can improve our understanding of FcγRs on macrophages; the established approach will facilitate further research on physiological activities involved in antibody-based immunotherapy.
Fc 受体（FcγR）广泛表达于效应细胞（如 NK 细胞、巨噬细胞），在癌症临床免疫治疗中发挥着重要作用。FcγR 与与靶细胞结合的抗体的 Fc 部分结合，可以激活抗体依赖的细胞介导的细胞毒性（ADCC）杀伤机制，导致靶细胞溶解。在这项工作中，我们使用原子力显微镜（AFM）在水相环境中观察单个巨噬细胞的细胞超微结构，并测量 FcγR 的生物物理性质（亲和力和分布）。AFM 成像用于获取巨噬细胞的形貌，揭示了纳米级细胞精细结构。对于分子相互作用识别，抗体分子通过异双功能聚乙二醇（PEG）交联剂连接到 AFM 探针上。通过 AFM 单分子力谱法，在单个巨噬细胞上定量测量 FcγR 的结合亲和力。使用粘附力映射方法定位 FcγR，揭示了巨噬细胞局部区域 FcγR 的纳米级分布。实验结果可以提高我们对巨噬细胞上 FcγR 的认识；所建立的方法将有助于进一步研究抗体免疫治疗中涉及的生理活动。
