Confocal reference free traction force microscopy

Abstract

The mechanical wiring between cells and their surroundings is fundamental to the regulation of complex biological processes during tissue development, repair or pathology. Traction force microscopy (TFM) enables determination of the actuating forces. Despite progress, important limitations with intrusion effects in low resolution 2D pillar-based methods or disruptive intermediate steps of cell removal and substrate relaxation in high-resolution continuum TFM methods need to be overcome. Here we introduce a novel method allowing a one-shot (live) acquisition of continuous in- and out-of-plane traction fields with high sensitivity. The method is based on electrohydrodynamic nanodrip-printing of quantum dots into confocal monocrystalline arrays, rendering individually identifiable point light sources on compliant substrates. We demonstrate the undisrupted reference-free acquisition and quantification of high-resolution continuous force fields, and the simultaneous capability of this method to correlatively overlap traction forces with spatial localization of proteins revealed using immunofluorescence methods.

Microfluidic traction force microscopy to study mechanotransduction in angiogenesis

“…Challenges, such as the tracking of markers within three dimensions, and the consideration of anisotropic substrate properties, could be overcome with the implementation of improved computational algorithms. These can provide even greater resolution, and aid the development of reference‐free approaches by removing the need to detach cells to obtain the reference location of trackers . Increased accuracy is vital to look deeper at the subcellular level, to study the coordination of cytoskeletal structure and sites of mechanoreceptor activation and traction force generation …”

Superresolved microparticle traction force microscopy reveals subcellular force patterns in immune cell-target interactions

“…This simplifies experimental procedures, is less disruptive to the sample, and will allow larger throughput studies. 35 We have focused on immune cell-target interactions and demonstrated detailed quantification of surprising patterns of cellular force exertion during these processes.…”

Persistent cell motility requires transcriptional feedback of cytoskeletal — focal adhesion equilibrium by YAP/TAZ

“…YAP/TAZ subcellular localization is regulated by cell-cell interactions through the Hippo pathway 19 and by cell-matrix interactions through cytoskeletal tension associated with cell spreading 11,12,20 .…”

“…C), had no effect on wound closure in either control or YAP/TAZ-depleted cells (p > .30; Supplementary Figure 2j). Intracellular mechanics dynamically respond to extracellular stimuli like contact inhibition release during migration [22][23][24] and drive YAP/TAZ nuclear localization 11,20 . Cellular resistance to perpendicular compressive forces at the cell apex is mediated by the tensile actomyosin network.…”