Evaluation of fluorophores for optimal performance in localization-based super-resolution imaging
By Graham T Dempsey, J. Vaughan, Kok Hao Chen, M. Bates, X. Zhuang. Nature Methods, 2011
1,278 citations recorded by the source corpus.
One approach to super-resolution fluorescence imaging uses sequential activation and localization of individual fluorophores to achieve high spatial resolution. Essential to this technique is the choice of fluorescent probes; the properties of the probes, including photons per switching event, on-off duty cycle, photostability and number of switching cycles, largely dictate the quality of super-resolution images. Although many probes have been reported, a systematic characterization of the properties of these probes and their impact on super-resolution image quality has been described in only a few cases. Here we quantitatively characterized the switching properties of 26 organic dyes and directly related these properties to the quality of super-resolution images. This analysis provides guidelines for characterization of super-resolution probes and a resource for selecting probes based on performance. Our evaluation identified several photoswitchable dyes with good to excellent performance in four independent spectral ranges, with which we demonstrated low–cross-talk, four-color super-resolution imaging.
Subfields: Biomedical Engineering, Biophysics
View Paper: https://www.semanticscholar.org/paper/40a63a70ab55698eb029866a44458fbca32bdad4
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In Biophysics
Citations in the corpus
- Nuclear pores as versatile reference standards for quantitative superresolution microscopy (2019, positive): “We confirmed that AF647, compared to the MEA buffer, shows increased brightness and number of localizations per fluorophore in BME 34 , reduced brightness and number of localizations per fluorophore in sulfite buffer 35 , and no substantial change in D 2 O 36 .”
- A general method to improve fluorophores for live-cell and single-molecule microscopy (2015, background): “Commercial compounds for spectroscopy were of the highest quality available and obtained from the following sources: Exciton ( 1 ); Anaspec ( 2 ); Promega ( 10 ); Sigma–Aldrich ( 11 , 13 ); Life Technologies ( 17 ); AAT Bioquest ( 23 ); New England BioLabs ( 30 ).”