High-throughput characterization of protein-protein interactions and assembly using differential dynamic microscopy

Understanding protein assembly requires comprehensive characterization of protein-protein interactions and assembly structures across a range of conditions spanning their native environment to technological applications, including complex, time-evolving milieu both in vivo and in vitro. However, conventional biophysical probes often require homogeneous or dilute samples, invasive sample preparation, or large material requirements. By contrast, recent studies have demonstrated how high-speed video microscopy of protein solutions can be combined with differential dynamic microscopy (DDM) to extract the hydrodynamic size, virial coefficient, and phase behavior associated with protein-protein interactions akin to conventional light scattering. By relying solely on concentration fluctuations of species below the diffraction limit, protein DDM can be performed on a range of systems without the need for complex labeling or super resolution microscopy to resolve protein structure and dynamics. Here, we demonstrate how protein DDM can be adapted for high-throughput measurements and analysis, providing insight into the fundamental interactions driving assembly of an intrinsically disordered protein. The results show promise for developing intensified, high-throughput measurements of scarce biomolecular materials to aid discovery and design of natural and engineered protein assemblies.