Cy5B-trisulfo, NHS
Biological Activity
Key information: Cy5B-trisulfo, NHS is a far-red fluorescent dye; supplied with an NHS ester reactive group for the labeling of primary amines. Particularly useful for smFRET experiments to reduce blinking and bleaching artefacts and maintain correct mean FRET efficiencies.Application: FRET, smFRET, FLIM, TIRF, and super resolution microscopy (SRM) techniques, including dSTORM. Provides enhanced spatial resolution in dSTORM experiments. Does not provide efficient blinking in conventional high-thiol dSTORM buffers; instead, sodium borohydride (NaBH4) addition is required following which SMLM experiments can be performed in PBS.
Properties and Photophysical Data: Excitation and emission maxima (λ) are 669 nm and 685 nm, respectively; extinction coefficient = 241,000 M-1cm-1; quantum yield = 0.4; fluorescence lifetime = 1.85 ns.
We suggest making up a 10 mM stock solution of this product in anhydrous DMSO.
For more information on how Cy5B-trisulfo, NHS may be used, see our protocol: Conjugation Protocol for Amine Reactive Dyes.
Scientific Data
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Application of Cy5B-trisulfo, NHS in COS7. FLIM composite image of a COS7 cell immunostained with mouse anti-beta -tubulin primary and Cyanine 5 labelled secondary (green), and rabbit anti-clathrin primary plus Cy5B-trisulfo, NHS labelled secondary (yellow-red). Scale bars, 5 μm and 1 μm (insets). Image kindly provided by Dr. Martin Schnermann, Center for Cancer Research, National Cancer Institute.
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Application of Cy5B-trisulfo, NHS in microtubules. Single-molecule localization microscopy (SMLM) images obtained using TIRF microscopy of microtubules immunolabeled with Cy5B-trisulfo, NHS labeled anti-tubulin antibody in 0.1% NaBH4/PBS upon irradiation with 640 nm (2.5 kWcm-2 ) using an integration time of 20 ms (total acquisition time 10 min). Scale bars, 3 μm and 1 μm (insets). Fourier ring correlation (FRC) analysis was used to estimate the indicated spatial resolutions. Image kindly provided by Dr. Martin Schnermann, Center for Cancer Research, National Cancer Institute.
Optical Data for Cy5B-trisulfo, NHS
| Emission Color | Far Red |
|---|---|
| λabs | 669 nm |
| λem | 685 nm |
| Extinction Coefficient (ε) | 241,000 M-1cm-1 |
| Quantum Yield (φ) | 0.4 |
| Closest Laser line | 640 nm |
| Reactive Group | NHS ester |
| Reactivity | Primary amines |
| Application | SMLM, FRET, smFRET, FLIM, dSTORM |
Plan Your Experiments
Use our spectra viewer to interactively plan your experiments, assessing multiplexing options. View the excitation and emission spectra for our fluorescent dye range and other commonly used dyes.
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Technical Data
The technical data provided above is for guidance only.
For batch specific data refer to the Certificate of Analysis.
Tocris products are intended for laboratory research use only, unless stated otherwise.
Additional Information
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