Fabrication of Nanomechanical Biosensors to Map 3D Surface Strain in Live Cells and Tissue

Abstract

This protocol details the step-by-step fabrication process to create an extracellular matrix (ECM) protein-based nanomechanical biosensor (NMBS) via an adaptation of the surface-initiated assembly (SIA) procedure1. The NMBS is a fluorescently labeled, ultra-thin fibronectin lattice-mesh with spatial resolution tailored by adjusting the width and spacing of the lattice from 2-100 µm. By applying the NMBS to the surface of cells and tissues one can directly measure deformation from subcellular to tissue length-scales. The procedure outlined here covers all aspects of NMBS construction from pattern design, photolithography for mold creation, casting of polydimethylsiloxane (PDMS) silicone stamps, conjugation of fluorescent dye to fibronectin, microcontact printing of fibronectin, creation of gelatin carriers2, transfer of the NMBS to gelatin carriers, and the use of gelatin carriers for application of the NMBS onto cells and tissue. The protocol can be broken down into three phases: material preparation (2 days), NMBS patterning (4 hours), and transfer to cell or tissue (1 hour). Material preparation and NMBS patterning can be performed ahead of time and the patterned NMBS can be stored for up to 1 year prior to use.