Self-Quenched Fluorophore-DNA Labels for Super-Resolution Fluorescence Microscopy
Laurell F. Kessler, Ashwin Balakrishnan, Tanja Menche, Dongni Wang, Yunqing Li, Maximilian Mantel, Marius Glogger, Marina S. Dietz, Mike Heilemann
Abstract
Protein labeling through transient and repetitive hybridization of short, fluorophore-labeled DNA oligonucleotides has become widely applied in various optical super-resolution microscopy methods. The main advantages are multitarget imaging and molecular quantification. A challenge is the high background signal originating from the presence of unbound fluorophore-DNA labels in solution. Here, we report the self-quenching of fluorophore dimers conjugated to DNA oligonucleotides as a general concept to reduce the fluorescence background. Upon hybridization, the fluorescence signals of both fluorophores are restored. We expand the toolbox of fluorophores suitable for self-quenching and report their spectra and hybridization equilibria. We apply self-quenched fluorophore-DNA labels to stimulated emission depletion microscopy and single-molecule localization microscopy and report improved imaging performances.