An MST-based assay reveals new binding preferences of IFIT1 for canonically and noncanonically capped RNAs

Workflow for the determination of K_D values for the IFIT1-capped RNA₇₀ interaction using an MST-based binding assay. Fluorescently labeled IFIT1 was mixed with RNA ligand at 16 different concentrations, equilibrated for 15 min, followed by the analysis by MST. The observed changes in fluorescence intensity were plotted as a function of ligand concentration and a theoretical binding curve was fitted to determine the dissociation constants (K_D).