Quantifying binding kinetics with fluorescence microscopy.

Abstract

Many processes crucial to life are regulated by the binding and unbinding of key molecules, such as target molecules to allosteric protein sites or transcription factors to gene loci. Capturing such binding events in living systems is often accomplished using fluorescence based methods. However, accurately determining assembly and disassembly rates from fluorescence data is challenging on account of the photophysical behavior of the fluorescent labels such as blinking and photobleaching. Here we present a Bayesian method that can simultaneously learn binding and unbinding rates while correcting for photophysical artifacts. We show that our method is robust across low light regimes relevant to in vivo experiments. We benchmark our method on both simulated and experimental data.