Live-cell monochromatic dual-label sub-diffraction microscopy by mt-pcSOFI

Abstract

We expand photochromic super-resolution optical fluctuation imaging (pcSOFI) to monochromatic dual-channel sub-diffraction microscopy. Multi-tau (mt-)pcSOFI unmixes spectrally identical reversibly switchable fluorescent proteins (RSFPs) based on their blinking kinetics. We show that mt-pcSOFI can be used to simultaneously image two structures in living cells with existing RSFPs and the newly developed ffDronpa-F.