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Nanoscale Research Letters

Ground-State Depletion Nanoscopy of Nitrogen-Vacancy Centres in Nanodiamonds

Jelle Storterboom, Martina Barbiero, Stefania Castelletto, Miṅ Gu

Published Mar 10, 2021• 10 citations
Publisher

Abstract

The negatively charged nitrogen-vacancy ([Formula: see text]) centre in nanodiamonds (NDs) has been recently studied for applications in cellular imaging due to its better photo-stability and biocompatibility if compared to other fluorophores. Super-resolution imaging achieving 20-nm resolution of [Formula: see text] in NDs has been proved over the years using sub-diffraction limited imaging approaches such as single molecule stochastic localisation microscopy and stimulated emission depletion microscopy. Here we show the first demonstration of ground-state depletion (GSD) nanoscopy of these centres in NDs using three beams, a probe beam, a depletion beam and a reset beam. The depletion beam at 638 nm forces the [Formula: see text] centres to the metastable dark state everywhere but in the local minimum, while a Gaussian beam at 594 nm probes the [Formula: see text] centres and a 488-nm reset beam is used to repopulate the excited state. Super-resolution imaging of a single [Formula: see text] centre with a full width at half maximum of 36 nm is demonstrated, and two adjacent [Formula: see text] centres separated by 72 nm are resolved. GSD microscopy is here applied to [Formula: see text] in NDs with a much lower optical power compared to bulk diamond. This work demonstrates the need to control the NDs nitrogen concentration to tailor their application in super-resolution imaging methods and paves the way for studies of [Formula: see text] in NDs' nanoscale interactions.

DOI
10.1186/s11671-021-03503-4

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The negatively charged nitrogen-vacancy ([Formula: see text]) centre in nanodiamonds (NDs) has been recently studied for applications in cellular imaging due to its better photo-stability and biocompatibility if compared to other fluorophores. Super-resolution imaging achieving 20-nm resolution of [Formula: see text] in NDs has been proved over the years using sub-diffraction limited imaging approaches such as single molecule stochastic localisation microscopy and stimulated emission depletion microscopy. Here we show the first demonstration of ground-state depletion (GSD) nanoscopy of these centres in NDs using three beams, a probe beam, a depletion beam and a reset beam. The depletion beam at 638 nm forces the [Formula: see text] centres to the metastable dark state everywhere but in the local minimum, while a Gaussian beam at 594 nm probes the [Formula: see text] centres and a 488-nm reset beam is used to repopulate the excited state. Super-resolution imaging of a single [Formula: see text] centre with a full width at half maximum of 36 nm is demonstrated, and two adjacent [Formula: see text] centres separated by 72 nm are resolved. GSD microscopy is here applied to [Formula: see text] in NDs with a much lower optical power compared to bulk diamond. This work demonstrates the need to control the NDs nitrogen concentration to tailor their application in super-resolution imaging methods and paves the way for studies of [Formula: see text] in NDs' nanoscale interactions.
Nitrogen-vacancy (NV) color centers in nanodiamonds are highly promising for bioimaging and sensing. However, resolving individual NV centers within nanodiamond particles and the controlled addressing and readout of their spin state has remained a major challenge. Spatially stochastic super-resolution techniques cannot provide this capability in principle, whereas coordinate-controlled super-resolution imaging methods, like stimulated emission depletion (STED) microscopy, have been predicted to fail in nanodiamonds. Here we show that, contrary to these predictions, STED can resolve single NV centers in 40-250 nm sized nanodiamonds with a resolution of ≈10 nm. Even multiple adjacent NVs located in single nanodiamonds can be imaged individually down to relative distances of ≈15 nm. Far-field optical super-resolution of NVs inside nanodiamonds is highly relevant for bioimaging applications of these fluorescent nanolabels. The targeted addressing and readout of individual NV(-) spins inside nanodiamonds by STED should also be of high significance for quantum sensing and information applications.
Due to their exceptional optical and magnetic properties, negatively charged nitrogen-vacancy (NV−) centers in nanodiamonds (NDs) have been identified as an indispensable tool for imaging, sensing and quantum bit manipulation. The investigation of the emission behaviors of single NV− centers at the nanoscale is of paramount importance and underpins their use in applications ranging from quantum computation to super-resolution imaging. Here, we report on a spin-manipulated nanoscopy method for nanoscale resolutions of the collectively blinking NV− centers confined within the diffraction-limited region. Using wide-field localization microscopy combined with nanoscale spin manipulation and the assistance of a microwave source tuned to the optically detected magnetic resonance (ODMR) frequency, we discovered that two collectively blinking NV− centers can be resolved. Furthermore, when the collective emitters possess the same ground state spin transition frequency, the proposed method allows the resolving of each single NV− center via an external magnetic field used to split the resonant dips. In spin manipulation, the three-level blinking dynamics provide the means to resolve two NV− centers separated by distances of 23 nm. The method presented here offers a new platform for studying and imaging spin-related quantum interactions at the nanoscale with super-resolution techniques. A new spin-manipulated nanoscopy method can resolve collectively blinking negatively charged nitrogen-vacancy centers (NVCs) on a nanoscale. The exceptional optical and magnetic properties of negatively charged nitrogen-vacancy centers in nanodiamonds make them attractive as fluorescent biomarkers in imaging and sensing applications. By combining wide-field localization microscopy with nanoscale spin manipulation by a microwave signal, Min Gu and colleagues from RMIT University and Swinburne University of Technology in Australia have achieved super-resolution imaging of the blinking of multiple, closely spaced negatively charged NVCs. The nanoscopy approach can resolve NVCs as small as 42 nanometers and separated by as little as 23 nanometers. Two close NVCs often appear to emit coupled fluorescence as they exhibit a collective behavior. The method offers a new way to study and image spin-related quantum interactions at the nanoscale.
We investigated the suitability of nanodiamonds (NDs) with nitrogen-vacancy (NV) centres as samples for resolution tests in stimulated emission depletion (STED) microscopy. The nanodiamonds were prepared using three different methods including additional proton irradiation to enhance NV centre density. This high-density sample exhibited stronger photoluminescence intensity, greater fluorescence stability, and higher image contrast under STED conditions. All samples showed strong resistance to damage from high STED laser power. Our results highlight the robustness and effectiveness of NDs with NV centres as reliable standards for evaluating STED microscope resolution.
Stimulated emission depletion (STED) and structured illumination (SIM) are two commonly used techniques for super-resolution imaging. However, the performance of these two techniques has never been quantitatively compared side-by-side. Taking advantage of the non-photobleaching characteristic of NV centres in fluorescent nanodiamond (FND), we performed a comparative study for the resolution of these two methods with 35 nm FNDs at the single particle level, as well as with FND grown in bulk diamond material. Results show that STED provides more structural details, whereas SIM provides a larger field of view with a higher imaging speed. SIM may induce deconvolution smooth and orientational artifacts during its post-processing.
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.