Jana M. Say

Therapeutic Inflammatory Monocyte Modulation Using Immune-Modifying Microparticles

Negatively charged immune-modifying microparticles bind to the scavenger receptor MARCO on inflammatory monocytes, resulting in their apoptosis and reduced inflammatory damage in a range of diseases. A New Frontier in Immune Modulation Inflammatory monocytes markedly potentiate the immune pathology observed in many diseases, yet no therapy exists that specifically inhibits these cells. The therapeutic accessibility of monocytes in the bloodstream and their inherent propensity to engulf particulate material suggest that highly negatively charged microparticles might provide a readily translatable solution to this problem. These microparticles, referred to as immune-modifying microparticles (IMPs), may be derived from numerous compounds, including the biodegradable polymer poly(lactic-co-glycolic acid) (PLGA-IMP), already used in humans for inter alia dissolvable sutures. Getts et al. now show that upon infusion, IMPs bind to a receptor with a positive domain on inflammatory monocytes, resulting in monocyte sequestration in the spleen and apoptosis through a similar pathway observed for senescing leukocytes. This safe monocyte clearance pathway culminated in substantially reduced inflammatory tissue damage in mouse models of West Nile virus encephalitis, experimental autoimmune encephalomyelitis, peritonitis, colitis, and myocardial infarction. Together, the data suggest that IMPs could transform the treatment of acute inflammation. Indeed, phase 1/2 testing is planned to begin in 2014, with rapid translation supported by the availability of clinical-grade PLGA. Inflammatory monocyte-derived effector cells play an important role in the pathogenesis of numerous inflammatory diseases. However, no treatment option exists that is capable of modulating these cells specifically. We show that infused negatively charged, immune-modifying microparticles (IMPs), derived from polystyrene, microdiamonds, or biodegradable poly(lactic-co-glycolic) acid, were taken up by inflammatory monocytes, in an opsonin-independent fashion, via the macrophage receptor with collagenous structure (MARCO). Subsequently, these monocytes no longer trafficked to sites of inflammation; rather, IMP infusion caused their sequestration in the spleen through apoptotic cell clearance mechanisms and, ultimately, caspase-3–mediated apoptosis. Administration of IMPs in mouse models of myocardial infarction, experimental autoimmune encephalomyelitis, dextran sodium sulfate–induced colitis, thioglycollate-induced peritonitis, and lethal flavivirus encephalitis markedly reduced monocyte accumulation at inflammatory foci, reduced disease symptoms, and promoted tissue repair. Together, these data highlight the intricate interplay between scavenger receptors, the spleen, and inflammatory monocyte function and support the translation of IMPs for therapeutic use in diseases caused or potentiated by inflammatory monocytes.

Three-dimensional optical manipulation of a single electron spin

Nitrogen vacancy (NV) centres in diamond are promising elemental blocks for quantum optics, spin-based quantum information processing and high-resolution sensing. However, fully exploiting the capabilities of these NV centres requires suitable strategies to accurately manipulate them. Here, we use optical tweezers as a tool to achieve deterministic trapping and three-dimensional spatial manipulation of individual nanodiamonds hosting a single NV spin. Remarkably, we find that the NV axis is nearly fixed inside the trap and can be controlled in situ by adjusting the polarization of the trapping light. By combining this unique spatial and angular control with coherent manipulation of the NV spin and fluorescence lifetime measurements near an integrated photonic system, we demonstrate individual optically trapped NV centres as a novel route for both three-dimensional vectorial magnetometry and sensing of the local density of optical states.

Effect of labeling with Iron oxide particles or nanodiamonds on the functionality of adipose-derived mesenchymal stem cells

Stem cells are increasingly the focus of translational research as well as having emerging roles in human cellular therapy. To support these uses there is a need for improved methods for in vivo cell localization and tracking. In this study, we examined the effects of cell labeling on the in vitro functionality of human adipose-derived mesenchymal stem cells. Our results provide a basis for future in vivo studies investigating implanted cell fate and longevity. In particular, we investigated the effects of two different particles: micron-sized (∼0.9 µm) fluorescently labeled (Dragon Green) superparamagnetic iron oxide particles (M-SPIO particles); and, carboxylated nanodiamonds of ∼0.25 µm in size. The effects of labeling on the functionality of adipose-derived MSCs were assessed by in vitro morphology, osteogenic and adipogenic differentiation potential, CD marker expression, cytokine secretion profiling and quantitative proteomics of the intra-cellular proteome. The differentiation and CD marker assays for stem-like functionality were not altered upon label incorporation and no secreted or intra-cellular protein changes indicative of stress or toxicity were detected. These in vitro results indicate that the M-SPIO particles and nanodiamonds investigated in this study are biocompatible with MSCs and therefore would be suitable labels for cell localization and tracking in vivo.

Luminescent nanodiamonds for biomedical applications

In recent years, nanodiamonds have emerged from primarily an industrial and mechanical applications base, to potentially underpinning sophisticated new technologies in biomedical and quantum science. Nanodiamonds are relatively inexpensive, biocompatible, easy to surface functionalise and optically stable. This combination of physical properties are ideally suited to biological applications, including intracellular labelling and tracking, extracellular drug delivery and adsorptive detection of bioactive molecules. Here we describe some of the methods and challenges for processing nanodiamond materials, detection schemes and some of the leading applications currently under investigation.

Emission and nonradiative decay of nanodiamond NV centers in a low refractive index environment

The nitrogen vacancy (NV) center is the most widely studied single optical defect in diamond with great potential for applications in quantum technologies. Development of practical single-photon devices requires an understanding of the emission under a range of conditions and environments. In this work, we study the properties of a single NV center in nanodiamonds embedded in an air-like silica aerogel environment which provides a new domain for probing the emission behavior of NV centers in nanoscale environments. In this arrangement, the emission rate is governed primarily by the diamond crystal lattice with negligible contribution from the surrounding environment. This is in contrast to the conventional approach of studying nanodiamonds on a glass coverslip. We observe an increase in the mean lifetime due to the absence of a dielectric interface near the emitting dipoles and a distribution arising from the irregularities in the nanodiamond geometry. Our approach results in the estimation of the mean quantum efficiency (~0.7) of the nanodiamond NV emitters.

A spectroscopic and carbon-isotope study of mixed-habit diamonds: Impurity characteristics and growth environment

Abstract Mixed-habit diamonds have experienced periods of growth where they were bounded by two surface forms at the same time. Such diamonds are relatively rare and therefore under-investigated. Under certain physical and chemical conditions, smooth octahedral faces grow concurrently with rough, hummocky cuboid faces. However, the specific conditions that cause this type of growth are unknown. Here we present a large array of spectroscopic data in an attempt to investigate the impurity and carbon-isotope characteristics, as well as growth conditions, of 13 large (>6 mm diameter) plates cut from mixed-habit diamonds. The diamonds all generally have high nitrogen concentrations (>1400 ppm), with the octahedral sectors enriched by 127-143% compared to their contemporary cuboid sectors. Levels of nitrogen aggregation are generally low (2-23% IaB) with no significant difference between sectors. IR-active hydrogen features are predominantly found in the cuboid sectors with only very small bands in the octahedral sectors. Platelet characteristics are variable; only one sample shows a large B′ band intensity in the octahedral sector, with no platelets occurring in the cuboid sector. Other samples either show a small B′ band in both sectors, or just in the cuboid sector, or none at all. These data support a model that shows the concentration-adjusted aggregation rate of nitrogen to be the same in both sectors, whereas the subsequent platelet development is reduced in the cuboid sectors. This is because the interstitial carbon atoms have interacted with disk-crack-like defects only found in cuboid sectors, which in turn reduces their chances of aggregating to form platelets. These disk-crack-like defects are also thought to be the most likely site for the IR-active hydrogen features and they maybe intrinsic to cuboid growth in mixed-habit diamonds. When they are graphitized, as they are in all of the diamonds in this study, this may reflect a heating event prior to volcanic exhumation. Spectroscopic analysis of the green cathodoluminescence exhibited by all of the diamonds shows nickel centers to be present in only the cuboid sectors. Carbon isotope data, obtained by secondary ion mass spectrometry, show very little variation in seven of the diamonds. The total range of 217 analyses is -7.94 to -9.61 (±0.15)‰, and the largest variation in a single stone is 0.98‰. No fractionation in carbon isotopes is seen between octahedral and cuboid sectors at the same growth horizon. These data suggest that the source fluid chemistry, as well as pressure, temperature, and oxygen fugacity were very stable over time, allowing such large volumes of mixed-habit growth to occur. The high concentration of impurities, namely nitrogen and hydrogen, is probably the critical factor required to cause mixed-habit growth. The impurity and isotopic data fall in line with previous modeling based on diamond growth from reduced carbonates with the loss of a 13C-enriched CO2 component.

A modular design of low-background bioassays based on a high-affinity molecular pair barstar:barnase

High‐affinity molecular pairs provide a convenient and flexible modular base for the design of molecular probes and protein/antigen assays. Specificity and sensitivity performance indicators of a bioassay critically depend on the dissociation constant (KD) of the molecular pair, with avidin:biotin being the state‐of‐the‐art molecular pair (KD ∼ 1 fM) used almost universally for applications in the fields of nanotechnology and proteomics. In this paper, we present an alternative high‐affinity protein pair, barstar:barnase (KD ∼ 10 fM), which addresses several shortfalls of the avidin:biotin system, including non‐negligible background due to the non‐specific binding. A quantitative assessment of the non‑specific binding carried out using a model assay revealed inherent irreproducibility of the [strept]avidin:biotin‐based assays, attributed to the avidin binding to solid phases, endogenous biotin molecules and serum proteins. On the other hand, the model assays assembled via a barstar:barnase protein linker proved to be immune to such non‐specific binding, showing good prospects for high‐sensitivity rare biomolecular event nanoproteomic assays.

Processing 15-nm Nanodiamonds Containing Nitrogen-vacancy Centres for Single-molecule FRET

Colour centres in nanodiamonds have many properties such as chemical and physical stability, biocompatibility, straightforward surface functionalisation as well as bright and stable photoluminescence, which make them attractive for biological applications. Here we examine the use of fluorescent nanodiamonds containing a single nitrogen-vacancy (NV) centre, as an alternative nano-label over conventional fluorophores. We describe a series of chemical treatments and air oxidation to reliably produce small (~15 nm) oxidised nanodiamonds suitable for applications in bioscience. We use Forster resonance energy transfer to measure the coupling efficiency from a single NV centre in a selected nanodiamond to an IRDye 800CW dye molecule absorbed onto the surface. Our single-molecule Forster resonance energy transfer analysis, based on fluorescence lifetime measurements, locates the position of the photostable NV centre deep within the core of the nanodiamond.

Nano-assembly of nanodiamonds by conjugation to actin filaments.

Fluorescent nanodiamonds (NDs) are remarkable objects. They possess unique mechanical and optical properties combined with high surface areas and controllable surface reactivity. They are non-toxic and hence suited for use in biological environments. NDs are also readily available and commercially inexpensive. Here, the exceptional capability of controlling and tailoring their surface chemistry is demonstrated. Small, bright diamond nanocrystals (size ˜30 nm) are conjugated to protein filaments of actin (length ˜3-7 µm). The conjugation to actin filaments is extremely selective and highly target-specific. These unique features, together with the relative simplicity of the conjugation-targeting method, make functionalised nanodiamonds a powerful and versatile platform in biomedicine and quantum nanotechnologies. Applications ranging from using NDs as superior biological markers to, potentially, developing novel bottom-up approaches for the fabrication of hybrid quantum devices that would bridge across the bio/solid-state interface are presented and discussed.

3D optical manipulation of a single electron spin

We optically trap individual nanodiamonds hosting a single Nitrogen Vacancy center and demonstrate a novel route for both non-invasive 3D vectorial magnetometry and sensing of the local density of states in a liquid environment.