Generation of biocompatible droplets for in vivo and in vitro measurement of cell-generated mechanical stresses

Here we describe a detailed protocol to produce biocompatible droplets that permit the measurement of mechanical stresses at cell and tissue scales. The droplets can be used as force transducers in vivo, ex vivo, and in vitro, to measure mechanical stresses in situ, in three dimensions and time. Versatile and modular droplet coatings using biotinylated molecules, such as ligands for specific adhesion receptors, enable the targeting of specific tissues or cells. Droplet sizes can be varied to measure forces at different scales (tissue and cell scales) and the range of measurable mechanical stresses ranges within approximately 0.3-100 kPa. The protocol described in this chapter is divided in three sections. First, we describe the generation and stabilization of biocompatible droplets. Next, we explain the steps necessary to functionalize the droplet surface. Finally, we describe how to characterize the mechanical properties of the droplets, so that they can be used as calibrated mechanical probes. The procedure to generate, stabilize, and functionalize the droplets is straightforward and can be completed in about 3 h with basic laboratory resources. The calibration of the droplet's mechanical properties to perform quantitative stress measurements is also straightforward, but requires the proper equipment to measure interfacial tension (such as a tensiometer). Calibrated droplets can be used to quantify cell-generated mechanical stresses by analyzing the tridimensional shape of the droplet.