High-Throughput Microrheology for the Assessment of Protein Gelation Kinetics

A high-throughput microrheological assay is employed to assess the gelation kinetics of a coiled-coil protein, Q, across a compositional space with varying ionic strengths and pH values. Two methods of passive microrheologymultiple particle tracking (MPT) and differential dynamic microscopy (DDM)are used to determine mean-squared displacements of tracer beads embedded in protein solutions with respect to lag time over a fixed period. MPT data was analyzed to determine gelation kinetics in a high-throughput, automatable manner by fitting relaxation exponents to sigmoidal curves and verifying with the more traditionally used time-cure superposition. DDM-determined gelation time was assessed as the last resolvable time, which we found to be on a similar scale to gelation times given by MPT. Both methods show distinct advantages with regard to being used in a high-throughput, automatable setup; DDM can serve as an effective initial screen for rapid gelation kinetics due to it requiring less user intervention and inputs, with MPT giving a more complete understanding of the entire gelation process. Using these methods, a clear optimum for rapid gelation was observed near the isoelectric point of Q and at higher ionic strengths over the compositional space studied.