Clot Imaging Using Photostable Nanodiamond.

While thrombosis is the leading cause of morbidity and mortality in the United States, an understanding of its triggers, progression, and response to anticoagulant therapy is lacking. Intravital fluorescence microscopy has advanced the study of thrombus formation by providing targeted, multi-color contrast. However, photodegradation of fluorophores limits the application in longitudinal studies (e.g., clot progression and/or dissolution). Fluorescent nanodiamond (FND) is a fluorophore which utilizes intrinsic fluorescence of chromogenic centers within and protected by the diamond crystalline lattice. Recent developments in diamond processing have allowed for the controlled production of nanodiamonds emitting in green or red. Here, the use of FND to label blood clots and/or clot lysis is demonstrated and compared to commonly used organic fluorophores. Model ex vivo clots were formed with incorporated labeled fibrinogen to allow imaging. FND was shown to match the morphology of organic fluorophore labels absent of photobleaching over time. The addition of tissue plasminogen activator (tPa) allowed visualization of the clot lysis stage, which is vital to studies of both DVT and pulmonary embolism resolution.

A Perspective on Fluorescent Nanodiamond Bioimaging.

The field of fluorescent nanodiamonds (FNDs) has advanced greatly over the past few years. Though historically limited primarily to red fluorescence, the wavelengths available for nanodiamonds have increased due to continuous technical advancement. This Review summarizes the strides made in the synthesis, functionalization, and application of FNDs to bioimaging. Highlights range from super-resolution microscopy, through cellular and whole animal imaging, up to constantly emerging fields including sensing and hyperpolarized magnetic resonance imaging.

Review Article: Synthesis, properties, and applications of fluorescent diamond particles.

Diamond particles containing color centers-fluorescent crystallographic defects embedded within the diamond lattice-outperform other classes of fluorophores by providing a combination of unmatched photostability, intriguing coupled magneto-optical properties, intrinsic biocompatibility, and outstanding mechanical and chemical robustness. This exceptional combination of properties positions fluorescent diamond particles as unique fluorophores with emerging applications in a variety of fields, including bioimaging, ultrasensitive metrology at the nanoscale, fluorescent tags in industrial applications, and even potentially as magnetic resonance imaging contrast agents. However, production of fluorescent nanodiamond (FND) is nontrivial, since it requires irradiation with high-energy particles to displace carbon atoms and create vacancies-a primary constituent in the majority color centers. In this review, centrally focused on material developments, major steps of FND production are discussed with emphasis on current challenges in the field and possible solutions. The authors demonstrate how the combination of fluorescent spectroscopy and electron paramagnetic resonance provides valuable insight into the types of radiation-induced defects formed and their evolution upon thermal annealing, thereby guiding FND performance optimization. A recent breakthrough process allowing for production of fluorescent diamond particles with vibrant blue, green, and red fluorescence is also discussed. Finally, the authors conclude with demonstrations of a few FND applications in the life science arena and in industry.

Targeting Fluorescent Nanodiamonds to Vascular Endothelial Growth Factor Receptors in Tumor.

The increased expression of vascular endothelial growth factor (VEGF) and its receptors is associated with angiogenesis in a growing tumor, presenting potential targets for tumor-selective imaging by way of targeted tracers. Though fluorescent tracers are used for targeted in vivo imaging, the lack of photostability and biocompatibility of many current fluorophores hinder their use in several applications involving long-term, continuous imaging. To address these problems, fluorescent nanodiamonds (FNDs), which exhibit infinite photostability and excellent biocompatibility, were explored as fluorophores in tracers for targeting VEGF receptors in growing tumors. To explore FND utility for imaging tumor VEGF receptors, we used click-chemistry to conjugate multiple copies of an engineered single-chain version of VEGF site-specifically derivatized with trans-cyclooctene (scVEGF-TCO) to 140 nm FND. The resulting targeting conjugates, FND-scVEGF, were then tested for functional activity of the scVEGF moieties through biochemical and tissue culture experiments and for selective tumor uptake in Balb/c mice with induced 4T1 carcinoma. We found that FND-scVEGF conjugates retain high affinity to VEGF receptors in cell culture experiments and observed preferential accumulation of FND-scVEGF in tumors relative to untargeted FND. Microspectroscopy provided unambiguous determination of FND within tissue by way of the unique spectral shape of nitrogen-vacancy induced fluorescence. These results validate and invite the use of targeted FND for diagnostic imaging and encourage further optimization of FND for fluorescence brightness.