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Nanodiamonds containing negatively charged nitrogen-vacancy (NV−) centers are versatile room-temperature quantum sensors in a growing field of research. Yet, knowledge regarding the NV− formation mechanism in very small particles is still limited. This study focuses on the formation of the smallest NV−-containing diamonds, 5 nm detonation nanodiamonds (DNDs). As a reliable method to quantify NV− centers in nanodiamonds, half-field signals in electron paramagnetic resonance (EPR) spectroscopy are recorded. By comparing the NV− concentration with a series of nanodiamonds from high-pressure high-temperature (HPHT) synthesis (10−100 nm), it is shown that the formation process in 5 nm DNDs is unique in several aspects. NV− centers in DNDs are already formed at the stage of electron irradiation, without the need for high-temperature annealing, an effect related to the very small particle size. Also, the NV− concentration (in atomic ratio) in 5 nm DNDs surpasses that of 20 nm-sized nanodiamonds, which contradicts the observation that the NV− concentration generally increases with particle size. This can be explained by the 10 times higher concentration of substitutional nitrogen atoms in the studied DNDs ([NS ≈ 1000 ppm]) compared to the HPHT nanodiamonds ([NS ≈ 100 ppm]). Upon electron irradiation at a fluence of 1.5 × 10^19 e−/cm2, DNDs show a 12.5-fold increment in the NV− concentration with no sign of saturation reaching 1 out of about 80 DNDs containing an NV− center. These findings can be of interest for the creation of defects in other very small semiconductor nanoparticles beyond NV-nanodiamonds as quantum sensors., The Journal of Physical Chemistry C, 126 (11), ISSN:1932-7455, ISSN:1932-7447

Detonation nanodiamonds (DNDs) are a class of very small and spherical diamond nanocrystals. They are used in polymer reinforcement materials or as drug delivery systems in the field of nanomedicine. Synthesized by detonation, only the final deaggregation step down to the single-digit nanometer size (< 10 nm) unfolds their full potential. Existing deaggregation methods mainly rely on mechanical forces, such as high-power sonication or beads milling. These techniques entail drawbacks such as contamination of the sample and the need for a specialized apparatus. In this paper, we report a purely chemical deaggregation method by simply combining oxidation in air followed by a boiling acid treatment, to produce highly stable single-digit DNDs in a suspension. The resulting DNDs are surface functionalized with carboxyl groups, the final boiling acid treatment removes primary metal contaminants such as magnesium, iron or copper and the nanoparticles remain dispersed over a wide pH range. Our method can be easily carried out in a standard chemistry laboratory with commonly available laboratory apparatus. This is a key step for many DND-based applications, ranging from material science to biological or medical applications.

Nanodiamonds containing negatively-charged Nitrogen-Vacancy (NV-) centers are versatile room-temperature quantum sensors in a growing field of research. Yet, knowledge regarding the formation mechanism in very small particles is still limited. Here, the study was focused on the formation of the smallest NV--containing diamonds, 5 nm detonation nanodiamonds (DNDs). As a reliable method to quantify NV- centers in nanodiamonds, half-field signals in electron paramagnetic resonance (EPR) spectroscopy are recorded. By comparing the NV- concentration in a series of nanodiamonds (5 - 100 nm), it was shown that the formation process in DNDs is unique in several aspects. NV- centers in DNDs are already formed during electron irradiation, without the need for high-temperature annealing. The effect was interpreted in terms of “self-annealing”, where size and type dependent effects enable vacancy migration at lower temperature. Although NV- concentration increases with particle size, the NV- concentration in NDs surpasses that of 20 nm-sized nanodiamonds. Using Monte-Carlo simulations, we show that the higher substitutional Nitrogen concentration compensates the vacancy loss induced by the large particle surface. Upon 1.5 × 1019 e-/cm2 electron irradiation, DNDs show a 12.5-fold NV- increment with no sign of saturation. These findings can be of interest for the creation of defects in other very small semiconductor nanoparticles.

Graphene oxide (GO) can form ultrapermeable and ultraselective membranes that are promising for various gas separation applications, including hydrogen purification. However, GO films lose their attractive separation properties in humid conditions. Here we show that incorporating positively charged nanodiamonds (ND+s) into GO nanolaminates leads to humidity-resistant, yet high-performing, membranes. While native GO membranes fail at a single run, the GO/ND+ composite retains up to ~90% of GO’s H2 selectivity against CO2 after several cycles under an aggressive humidity test. The addition of negatively charged ND to GO brought no such stabilization, suggesting that charge compensation acts as the main mechanism conferring humidity resistance, where ND+s neutralize the negative charge GO sheets. We observed a similar but inferior stabilization effect when positively charged polyhedral oligomeric silsesquioxane replaces ND+. The demonstrated material platform offers a solution for separating H2 gas from its usually humid mixtures generated from fossil fuel sources or water splitting.

We demonstrate room-temperature 13C hyperpolarization by dynamic nuclear polarization (DNP) using optically polarized triplet electron spins in two polycrystalline systems: pentacene-doped [carboxyl-13C] benzoic acid and microdia-monds containing NV−centers. For both samples, the integrated solid effect (ISE) is used to polarize the13C spin system in magnetic fields of 350-400 mT. In the benzoic acid sample, the13C spin polarization is enhanced up to 0.12% through directelectron-to-13C polarization transfer without performing dynamic1H polarization followed by 1H-13C cross polarization. In5addition, ISE has been successfully applied for the first time to polarize naturally abundant 13C spins in a microdiamond sample to 0.01%. To characterize the buildup of the13C polarization, we discuss the efficiencies of direct polarization transfer be-tween the electron and13C spins as well as that of13C–13C spin diffusion, examining various parameters which are beneficial or detrimental for successful bulk dynamic13C polarization.

Detonation nanodiamonds (DNDs) with sizes below 10 nm have attracted attention as single photon emitters with potential in many research fields from life sciences to quantum technologies. However, while nitrogen-vacancy (NV) color centers are found in nanodiamonds directly after the detonation synthesis without the need for irradiation or annealing, silicon-vacancy (SiV) color centers are not present in these pristine samples. Herein, SiV centers are created in DNDs by an annealing treatment up to 1100 °C in high vacuum. As silicon is not added, the precursor of the SiV centers must be pristine silicon impurities inside the nanodiamond lattice. A sharp emission line at the wavelength of 737 nm with a linewidth of 7.7 nm is observed in DNDs that are electron irradiated before annealing. This wavelength is consistent with the characteristic emission line of SiV centers and its linewidth is comparable with that in larger nanodiamonds created by chemical vapor deposition and subsequent milling. The average lifetime (0.4 ± 0.04 ns) of the fluorescence, which is in the range of reported lifetimes in nanodiamonds, also support that the observed emission peak are due to SiV centers in DNDs., Physica Status Solidi A, 218 (19), ISSN:1862-6300, ISSN:1862-6319, ISSN:0031-8965, ISSN:1521-396X

We demonstrate room-temperature 13C hyperpolarization by dynamic nuclear polarization (DNP) using optically polarized triplet electron spins in two polycrystalline systems: pentacene-doped [carboxyl-13C] benzoic acid and microdiamonds containing NV- centers. For both samples, the integrated solid effect (ISE) is used to polarize the 13C spin system in magnetic fields of 350–400 mT. In the benzoic acid sample, the 13C spin polarization is enhanced up to 0.12 % through direct electron-to-13C polarization transfer without performing dynamic 1H polarization followed by 1H-13C cross polarization. In addition, ISE has been successfully applied for the first time to polarize naturally abundant 13C spins in a microdiamond sample to 0.01 %. To characterize the buildup of the 13C polarization, we discuss the efficiencies of direct polarization transfer between the electron and 13C spins as well as that of 13C–13C spin diffusion, examining various parameters which are beneficial or detrimental for successful bulk dynamic 13C polarization.

Detonation nanodiamonds (DNDs) are a class of very small and spherical diamond nanocrystals. Used in polymer reinforcement materials, as drug delivery systems or as fluorescent biomarkers, only the final deaggregation step down to the single-digit nanometer size unfolds their full potential. All existing deaggregation methods rely on mechanical forces, such as high-power sonication or beads milling. We report a purely chemical deaggregation method by combining oxidation in air followed by a boiling acid treatment. Our DNDs are surface functionalized with carboxyl groups, the final boiling acid treatment removes primary metal contaminants and the nanoparticles remain dispersed over a wide pH range. Experiments can be easily carried out in a standard chemistry laboratory. This is a key step for many DND-based applications, ranging from material science to biological or medical applications and opens a way for inexpensive mass production on industrial scale.

Fluorescent nanodiamonds (FNDs) have been attracting much attention as promising therapeutic agents and probes for bioimaging and nanosensing. For their biological applications, several hydrophilizing methods to enhance FND colloidal stability have been developed to suppress their aggregation and the nonspecific adsorption to biomolecules in complex biomedical environments. However, these methods involve several complicated synthetic and purification steps, which prohibit the use of FNDs for bioapplications by biologists. In this study, we describe a simple one-pot FND hydrophilization method that comprises coating of the surface of the nanoparticles with COOH-terminated hyperbranched polyglycerol (HPG-COOH). HPG-COOH-coated FNDs (FND-HPG-COOHs) were found to exhibit excellent dispersibility under physiological conditions despite the thinness of the 5 nm HPG-COOH layer. Biotinylated FND-HPG-COOHs specifically captured avidin molecules in the absence of nonspecific protein adsorption. Moreover, we demonstrated that FND-HPG-COOHs conjugated with antibodies can be used to selectively target integrins in fixed HeLa cells. In addition, intracellular temperature changes were measured via optically detected magnetic resonance using FND-HPG-COOHs conjugated with mitochondrial localization signal peptides. Our one-pot synthetic method will encourage the broad use of FNDs among molecular and cellular biologists and pave the way for extensive biological and biomedical applications of FNDs.

Nanodiamonds have acknowledged growing attention due to their facile functionalization, stable fluorescence, low toxicity, and decent biocompatibility. However, despite advances applications of nanodiamonds in drug delivery, catalysis and bio-sensing, laser irradiation still limits long-time tracking of nanodiamonds in living cells due to phototoxicity. Here, using optical diffraction tomography, we performed quantitative morphological and biophysical analysis of living cells via endocytosis or electroporation of nanodiamonds (5 nm, 35 nm, and 100 nm) without the need for a fluorescence label. Optical diffraction tomography is an inexpensive and noninvasive microscopy technique, which images cells and subcellular structures as a function of their refractive index. The laser excitation power is much weaker than in the case of fluorescence microscopy, which reduces phototoxicity. Thanks to the very high refractive index of diamond, nanodiamonds in HeLa cells can be clearly discriminated from cellular structures using optical diffraction tomography. As an application, we show that aggregation and deaggregation of internalized nanodiamonds can be detected via changes in the refractive index distribution of the entire cell. Optical diffraction tomography successively images prevention of in-cell particle aggregation through polyglycerol coating of nanodiamonds. In the case of endocytosis, optical diffraction tomography shows deaggregation of nanodiamonds after a prolonged incubation time. Together, our findings implicate that refractive index measurements are a favorable tool to track nanodiamonds, without a fluorescent label, inside living cells. This could be useful to study real-time therapeutic or metabolic activities in living cells using very weak laser irradiation. Finally, the elaborate creation of fluorescent defects in nanodiamonds becomes redundant.

Computational protein design has advanced very rapidly over the last decade, but there remain few examples of artificial proteins with direct medical applications. This study describes a new artificial β-trefoil lectin that recognises Burkitt’s lymphoma cells, and which was designed with the intention of finding a basis for novel cancer treatments or diagnostics. The new protein, called “Mitsuba”, is based on the structure of the natural shellfish lectin MytiLec-1, a member of a small lectin family that uses unique sequence motifs to bind α-D-galactose. The three subdomains of MytiLec-1 each carry one galactose binding site, and the 149-residue protein forms a tight dimer in solution. Mitsuba (meaning “three-leaf” in Japanese) was created by symmetry constraining the structure of a MytiLec-1 subunit, resulting in a 150-residue sequence that contains three identical tandem repeats. Mitsuba-1 was expressed and crystallised to confirm the X-ray structure matches the predicted model. Mitsuba-1 recognises cancer cells that express globotriose (Galα(1,4)Galβ(1,4)Glc) on the surface, but the cytotoxicity is abolished.

Nanodiamonds containing negatively charged nitrogen-vacancy (NV-) centers are versatile nanosensors thanks to their optical and spin properties. While currently most fluorescent nanodiamonds in use have at least a size of a few tens of nanometers, the challenge lies in engineering the smallest nanodiamonds containing a single NV- defect. Such a tiny nanocrystal with a single NV- center is an "optical spin label" for biomolecules, which can be detected in a fluorescence microscope. In this paper, we address two key issues toward this goal using detonation nanodiamonds (DNDs) of 4-5 nm in size. The DND samples are treated first with electron irradiation to create more vacancies. With the aid of electron paramagnetic resonance (EPR) spectroscopy, we confirm a steady increase of negatively charged NV- centers with higher fluence. This leads to a 4 times higher concentration in NV- defects after irradiation with 2 MeV electrons at a fluence of 5 × 1018 e-/cm2. Interestingly, we observe that the annealing of DND does not increase the number of NV- centers, which is in contrast to bulk diamond and larger nanodiamonds. Since DNDs are strongly aggregated after the irradiation process, we apply a boiling acid treatment as a second step to fabricate monodisperse DNDs enriched in NV- centers. These are two important steps toward "optical spin labels" having a single-digit nanometer range size that could be used for bioimaging and nanosensing.

Introducing nanomaterial in interfacial polymerization (IP) system for nanofiltration (NF) membrane synthesis has witnessed a remarkable performance enhancement thus drawing intensive attention. However, the underlying mechanism for nanomaterial induced performance enhancement is still unclear due to the lack of study on nanoparticle dispersity and architecture at polymerization interface. Using nanodiamond (ND) as the example, this study demonstrates nanoparticle undergoes aggregation preferably at the reaction interface and the architecture of ND particles has a direct impact on membrane structure and performance. Through proactively controlling the aggregation extent while employing these ND clusters as the nano-template, the feature morphology of NF membrane is transformed from nodules to ridges at the nanoscale. Such transformation generates a significant augmentation of effective membrane area, leading to the increase of water permeance by 70%. With a low amount of nanodiamond addition (

Background: Nanodiamonds (NDs) provide a unique multitasking system for drug delivery and fluorescent imaging in biological environments. Owing to their quantum properties, NDs are expected to be employed as multifunctional probes in the future for the accurate visualization of biophysical parameters such as temperature and magnetic fields. However, the use of NDs for the selective targeting of the biomolecules of interest within a complicated biological system remains a challenge. One of the most promising solutions is the appropriate surface design of NDs based on organic chemistry and biochemistry. The engineered NDs have high biocompatibility and dispersibility in a biological environment and hence undergo cellular uptake through specific pathways. Scope of review: This review focuses on the selective targeting of NDs for biomedical and biophysical applications from the viewpoint of ND surface functionalizations and modifications. These pretreatments make possible the specific targeting of biomolecules of interest on or in a cell by NDs via a designed biochemical route. Major conclusions: The surface of a ND is covalently or noncovalently modified with silica, polymers, or biomolecules to reshape them, control their size, and enhance the colloidal stability and biomolecular selectivity toward biomolecules of interest. Electroporation, chemical treatment, injection, or endocytosis are the methods generally adopted to introduce a ND into living cells. The pathway, efficiency, and the cell viability depend on the selected method. General significance: In the biomedical field, surface modification facilitates specific delivery of a drug, leading to a higher therapeutic efficacy. In biophysical applications, surface modification paves the way for the accurate measurement of physical parameters to gain a better understanding of various cell functions.

Background Nanodiamonds (NDs) provide a unique multitasking system for drug delivery and fluorescent imaging in biological environments. Owing to their quantum properties, NDs are expected to be employed as multifunctional probes in the future for the accurate visualization of biophysical parameters such as temperature and magnetic fields. However, the use of NDs for the selective targeting of the biomolecules of interest within a complicated biological system remains a challenge. One of the most promising solutions is the appropriate surface design of NDs based on organic chemistry and biochemistry. The engineered NDs have high biocompatibility and dispersibility in a biological environment and hence undergo cellular uptake through specific pathways. Scope of review This review focuses on the selective targeting of NDs for biomedical and biophysical applications from the viewpoint of ND surface functionalizations and modifications. These pretreatments make possible the specific targeting of biomolecules of interest on or in a cell by NDs via a designed biochemical route. Major conclusions The surface of NDs is covalently or noncovalently modified with silica, polymers, or biomolecules to reshape them, control their size, and enhance the colloidal stability and biomolecular selectivity toward the biomolecules of interest. Electroporation, chemical treatment, injection, or endocytosis are the methods generally adopted to introduce NDs into living cells. The pathway, efficiency, and the cell viability depend on the selected method. General significance In the biomedical field, the surface modification facilitates specific delivery of a drug, leading to a higher therapeutic efficacy. In biophysical applications, the surface modification paves the way for the accurate measurement of physical parameters to gain a better understanding of various cell functions.

The development of sensors to estimate physical properties, and their temporal and spatial variation, has been a central driving force in scientific breakthroughs. In recent years, nanosensors based on quantum measurements, such as nitrogen-vacancy centres (NVCs) in nanodiamonds, have been attracting much attention as ultrastable, sensitive, accurate and versatile physical sensors for quantitative cellular measurements. However, the nanodiamonds currently available for use as sensors have diameters of several tens of nanometres, much larger than the usual size of a protein. Therefore, their actual applications remain limited. Here we show that NVCs in an aggregation of 5-nm-sized detonation-synthesized nanodiamond treated by Krüger’s surface reduction (termed DND-OH) retains the same characteristics as observed in larger diamonds. We show that the negative charge at the NVC are stabilized, have a relatively long T2 spin relaxation time of up to 4 μs, and are applicable to thermosensing, one-degree orientation determination and nanometric super-resolution imaging. Our results clearly demonstrate the significant potential of DND-OH as a physical sensor. Thus, DND-OH will raise new possibilities for spatiotemporal monitoring of live cells and dynamic biomolecules in individual cells at single-molecule resolution., Scientific Reports, 8 (1), ISSN:2045-2322

MytiLec is a lectin, isolated from bivalves, with cytotoxic activity against cancer cell lines that express globotriaosyl ceramide, Galα(1,4)Galβ(1,4)Glcα1-Cer, on the cell surface. Functional analysis shows that the protein binds to the disaccharide melibiose, Galα(1,6)Glc and the trisaccharide globotriose, Galα(1,4)Galβ(1,4)Glc. Recombinant MytiLec expressed in bacteria showed the same haemagglutinating and cytotoxic activity against Burkitt’s lymphoma (Raji) cells as the native form. The crystal structure has been determined to atomic resolution, in the presence and absence of ligands, showing the protein to be a member of the β-trefoil family, but with a mode of ligand binding unique to a small group of related trefoil lectins. Each of the three pseudo-equivalent binding sites within the monomer shows ligand binding and the protein forms a tight dimer in solution. An engineered monomer mutant lost all cytotoxic activity against Raji cells, but retained some haemagglutination activity, showing that the quaternary structure of the protein is important for its cellular effects.

MytiLec, a novel lectin isolated from the Mediterranean mussel (Mytilus galloprovincialis), shows strong binding affinity to globotriose (Gb3: Galα1-4Galβ1-4Glc). MytiLec revealed β-trefoil folding as also found in the ricin B-subunit type (R-type) lectin family, although the amino acid sequences were quite different. Classification of R-type lectin family members therefore needs to be based on conformation as well as on primary structure. MytiLec specifically killed Burkitt's lymphoma Ramos cells, which express Gb3. Fluorescein-labeling assay revealed that MytiLec was incorporated inside the cells. MytiLec treatment of Ramos cells resulted in activation of both classical MAPK/ extracellular signal-regulated kinase and extracellular signal-regulated kinase (MEK-ERK) and stress-activated (p38 kinase and JNK) Mitogen-activated protein kinases (MAPK) pathways. In the cells, MytiLec treatment triggered expression of tumor necrosis factor (TNF)-α (a ligand of death receptor-dependent apoptosis) and activation of mitochondria-controlling caspase-9 (initiator caspase) and caspase-3 (activator caspase). Experiments using the specific MEK inhibitor U0126 showed that MytiLec-induced phosphorylation of the MEK-ERK pathway up-regulated expression of the cyclin-dependent kinase inhibitor p21, leading to cell cycle arrest and TNF-α production. Activation of caspase-3 by MytiLec appeared to be regulated by multiple different pathways. Our findings, taken together, indicate that the novel R-type lectin MytiLec initiates programmed cell death of Burkitt’s lymphoma cells through multiple pathways (MAPK cascade, death receptor signaling, caspase activation) based on interaction of the lectin with Gb3-containing glycosphingolipid-enriched microdomains on the cell surface.