STED imaging of green fluorescent nanodiamonds containing nitrogen-vacancy-nitrogen centers

Abstract

Abstract: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. 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.

A simple empirical algorithm for optimising depletion power and resolution for dye and system specific STED imaging

“…In general, anything that alters absorption or scattering in the sample (or, like bead index, changes the lifetime of the probe) may affect P STED . Although there are many sites which give helpful information about the ‘best’ STED fluorophores (https://nanobiophotonics.mpibpc.mpg.de/dyes/, https://www.leica-microsystems.com/science-lab/quick-guide-to-sted-sample-preparation/, https://www.abberior-instruments.com/site/assets/files/2190/recommended_fluorescent_markers_for_sted_by_abberior_instruments-1.pdf) and new STED probes are being developed (Hense et al ., ; Maksim et al ., ; Rosales et al ., ; Butkevich et al ., ; Byrne et al ., ; Laporte & Psaltis, ; Butkevich et al ., ; Erdmann et al ., ; Tavernaro et al ., ), it is important to know how the dye is working locally on a given nanoscope.…”

Nanodiamonds and Their Applications in Cells

“…Besides their fluorescence, nanodiamonds exhibit a characteristic Raman peak at 1332 cm −1 , while a cell's Raman peak is around 2800–3200 cm −1 . This trait makes nanodiamond a promising probe for Raman mapping and imaging (Figure d) . Chao et al used Raman imaging to visualize how lysozyme interacts with bacteria .…”

Nanodiamond: A high impact nanomaterial

“…Complexes consisting of vacancies trapped by nitrogen atoms form different color centers. The nitrogen-vacancy (NV) defect, responsible for diamond's red/near-infrared fluorescence, and NVN color centers (H3 centers), with bright green fluorescence, are the optically active defects that have received the most consideration [19][20][21]27]. The NV center is a defect formed in diamond by one substitutional nitrogen atom and a vacancy, while a NVN center consists of a pair of N atoms adjacent to a vacancy.…”

“…The capability of optically detected magnetic resonance and stable photoluminescence of the NV À center make it an appealing candidate for nanoscale sensing, quantum computing, and bioimaging in vivo and in vitro applications [19,[36][37][38]. NV and NVN (H3) centers are also promising candidates for stimulated emission depletion (STED) imaging [27,39]. As they are present within the bulk of the diamond lattice and are protected from the surrounding environmental agents (such as strong chemicals), fluorescent defect centers in diamond offer infinitely stable fluorescence with no photo bleaching or blinking.…”

“…Aside from these opportunities, there are always continued fundamental topics which are hard to predict. Research involving the use of fluorescent nanodiamonds for super-resolution microscopy (in particular, STED) is an emerging topic which has exciting potential [27,39,40].…”