Protein binding kinetics quantification via coupled plasmonic-photonic resonance nanosensors in generic microplate reader

Abstract

Almost no analytical assays, either colorimetric or fluorescence assays, for generic microplate readers is capable of dynamic measurements of protein-protein binding or the quantification of kinetic association and dissociation constants of protein interactions. On the other hand, protein binding kinetics quantification can be uniquely done on special expensive surface plasmon resonance (SPR) sensing equipment. Here we report the integration of coupled plasmonic-photonic resonance nanosensors in standard 96-well plate format and by using which, for the very first time, the demonstration of label-free dynamic SPR-like protein binding measurement and kinetics quantification in a generic microplate reader. Our low-cost label-free nanosensor plate enables very sensitive detection of immobilized protein interactions based on the transmission optical density (OD) value changes at specific wavelengths measured in a generic microplate reader. The relative end-point OD value changes show a good linear response with protein concentrations (from 0.05 to 50 μg/ml). And the protein quantification in serum results are consistent with the concurrent hospital lab tests. Most importantly, the kinetic association and dissociation constants of protein interactions in our sensor plate wells are determined by time-lapse dynamic OD value measurement in the generic microplate reader. Enabled by our unique nanosensor plate, SPR-like measurement of protein binding kinetics is now available using generic microplate reader ubiquitous in many chemistry and biomedical research labs.