Droplet Based Measurements of Mechanical Forces and Material Properties, In Vivo and In Vitro

Abstract

The spatiotemporal patterning of physical stresses and mechanical properties inside living tissues ultimately guide morphogenesis, both during normal development and cancer progression. Here, we present various droplet-based techniques for measuring these physical quantities in biological systems, both in vivo and in vitro. We describe the methods for generating and functionalizing droplets, which can be placed in a tissue and imaged in 3D. We present an open source Matlab based computational tool recently made available to the community that performs automated 3D surface segmentation and curvature analysis from a volumetric image stack with typical errors in radial and curvature measurements less than 2 and 20 percent, respectively. By placing cell-sized fluorocarbon droplets within multicellular spheroids, we measure anisotropies in cell-generated stresses and quantify how these stresses change over time and space within the aggregates. Furthermore we present in situ calibrated measurements of cellular stresses inside the developing zebrafish tailbud (10-12 somite stage) obtained with ferrofluid droplets (30-50 microns in diameter). Ferrofluid droplets were also used to measure mechanical properties using magnetic actuation. In spheroids, stresses are observed to increase significantly over time while maintaining a weak spatial dependence. In vivo studies in the zebrafish tailbud, however, reveal no significant spatial dependence in the magnitude of cellular stresses, although patterning of the material properties is observed. This work highlights the potential for droplet based in situ measurements to elucidate mechanical phenomena underlying processes in developmental and cancer biology.