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DNA-Based Probes for Measuring Mechanical Forces in Cell-Cell Contacts: Application to B Cell Antigen Extraction from

Katelyn M Spillane1,2, Pavel Tolar3,4

  • 1Immune Receptor Activation Laboratory, The Francis Crick Institute, 1 Midland Road, London, NW1 1AT, UK.

Methods in Molecular Biology (Clifton, N.J.)
|February 2, 2018
PubMed

The production of antibodies requires the expansion and selection of high-affinity B cell clones. This process is initiated by antigen uptake through the B cell receptor (BCR), which recognizes and binds antigen displayed on the surface of an antigen-presenting cell (APC). To acquire the antigen, B cells use myosin contractility to physically pull BCR-antigen clusters from the APC membrane. These mechanical forces influence association and dissociation rates of BCR-antigen bonds, resulting in affinity-dependent acquisition of antigen by B cells. Mechanical regulation of B cell antigen acquisition from APCs remains poorly understood, although the recent development of DNA-based force sensors has enabled the measurement of mechanical forces generated in B cell-APC contacts. In this chapter, we describe a protocol to design, synthesize, and purify DNA-based force sensors to measure B cell antigen extraction forces using fluorescence microscopy.

Keywords:
Antigen internalizationB cell receptorB cellsCell mechanicsDNA force sensorDNA-protein conjugationFluorescence microscopyImmune synapse

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