Hannah C W McArthur, Anna T Bajur, Katelyn M Spillane
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引用次数: 0
摘要
B 细胞对抗原递呈细胞（APC）施加牵引力，以提取抗原进行内化。对 B 细胞受体（BCR）-抗原键施加拉力可促进抗原亲和力的判别和对 APC 物理特性的感知。在这里，我们介绍了一种制备抗原功能化 DNA 张力传感器的方法，这种传感器可用于量化 B 细胞免疫突触中力介导的抗原提取。我们介绍了如何将传感器附着在平面脂质双分子层上、量化其表面密度、使用它们刺激 B 细胞活化，以及通过荧光显微镜和图像分析来分析固定细胞中抗原提取的效率。这些技术应广泛适用于细胞-细胞接触中分子的力介导转移研究。
Quantifying force-mediated antigen extraction in the B cell immune synapse using DNA-based tension sensors.
B cells exert pulling forces against antigen-presenting cells (APCs) to extract antigens for internalization. The application of tugging forces on B cell receptor (BCR)-antigen bonds promotes discrimination of antigen affinities and sensing of APC physical properties. Here, we describe a protocol for preparing antigen-functionalized DNA tension sensors for quantifying force-mediated antigen extraction in the B cell immune synapse. We describe how to attach the sensors to planar lipid bilayers, quantify their surface density, use them to stimulate B cell activation, and analyze the efficiency of antigen extraction in fixed cells by fluorescence microscopy and image analysis. These techniques should be broadly applicable to studies of force-mediated transfer of molecules in cell-cell contacts.
期刊介绍：
For over fifty years, Methods in Cell Biology has helped researchers answer the question "What method should I use to study this cell biology problem?" Edited by leaders in the field, each thematic volume provides proven, state-of-art techniques, along with relevant historical background and theory, to aid researchers in efficient design and effective implementation of experimental methodologies. Over its many years of publication, Methods in Cell Biology has built up a deep library of biological methods to study model developmental organisms, organelles and cell systems, as well as comprehensive coverage of microscopy and other analytical approaches.
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