Quantifying force-mediated antigen extraction in the B cell immune synapse using DNA-based tension sensors.

Abstract

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.