biostudies-literatureKessler LFDeutsche ForschungsgemeinschaftGoethe-Universit?t Frankfurt am Main6751-6759https://www.ebi.ac.uk/biostudies/studies/S-EPMC11264260biostudies-literatureUnknown128(28)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 stimulatThe journal of physical chemistry. BSelf-Quenched Fluorophore-DNA Labels for Super-Resolution Fluorescence Microscopy.PMC11264260CRC 1177INST 161/1020-1 FUGGCRC 1507Mantel MLi YDietz MSMenche TWang DHeilemann MKessler LFBalakrishnan AGlogger MfalseSelf-Quenched Fluorophore-DNA Labels for Super-Resolution Fluorescence Microscopy.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 stimulat2024-01-01T00:00:00Z2024 Jul2025-06-01T01:15:48.184Z2025-06-01T01:15:48.184ZS-EPMC112642603895534610.1021/acs.jpcb.4c02065