Abstract
We report Stimulated Emission Depletion (STED) imaging of green fluorescent nanodiamonds containing Nitrogen-acancy-Nitrogen (NVN) centers with a resolution of 70 nm using a commercial microscope. Nanodiamonds have been demonstrated to have the potential to be excellent cellular biomarkers thanks to their low toxicity and nonbleaching fluorescence, and are especially appealing for superresolution imaging technique like STED microscopy. However, only red fluorescent nanodiamonds containing Nitrogen-Vacancy (NV) centers have been used with STED microscopy so far. The existence of only one color nonbleaching center limits the possible observations, for instance it complicates spatial correlation studies with STED. To provide a nonbleaching probe in a different color, we characterize here the optical properties of the NVN defect for STED imaging. We demonstrate STED imaging of the green fluorescent nanodiamonds containing NVN centers, opening the door for long term two-color STED observation. Furthermore we exemplify the use of green nanodiamonds as a second color nonbleaching STED biomarker by imaging 70 nm fluorescent crystals up taken into HeLa cells.
Highlights
Stimulated Emission Depletion (STED) microscopy is a superresolution imaging technique that has revolutionized optical microscopy in the last two decades
We report Stimulated Emission Depletion (STED) imaging of green fluorescent nanodiamonds containing Nitrogen-Vacancy-Nitrogen (NVN) centers with a resolution of 70 nm using a commercial microscope
We have demonstrated that the photophysical properties of green nanodiamonds (gNDs) allow their efficient use in STED microscopy
Summary
STED microscopy is a superresolution imaging technique that has revolutionized optical microscopy in the last two decades It results in breaking the diffraction limit of a fluorescence laser scanning microscope by depleting the fluorescence around the focus by stimulated emission. One solution is to develop dyes and fluorescent probes with less photobleaching In this sense color centers in diamond have an exceptional property: they do not exhibit photobleaching and are exceptionally photostable [6]. They are excellent luminescent sources for far field fluorescence nanoscopy by STED, and NV centers have been imaged in many STED microscopy studies [2,3,7]. In order to show the potential of green nanodiamonds (gNDs) as photostable biomarkers in fluorescence nanoscopy, we acquired superresolved images of 70 nm sized diamond particles uptaken into HeLa cells
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