Preparation and characterization of green fluorescent nanodiamonds for biological applications

Abstract

Green fluorescent nanodiamonds (denoted as gFNDs) have been produced and characterized for use as cellular markers and/or labels. The gFND particles, containing a high concentration of N–V–N (or H3) centers, were prepared by radiation damage of type Ia natural diamond nanocrystallites with either a home-built 40-keV He + beam or a research-grade 3-MeV H + beam, followed by thermal annealing at 800 °C. Prior to the irradiation treatment, infrared spectroscopy of C–N stretches (at 1282 cm − 1 ) of natural diamond single crystals revealed a nitrogen concentration in the range of 900 ppm. Irradiation and annealing of the specimens led to the emergence of a broad absorption band at ~ 470 nm, corresponding to the phonon sideband of the electronic transition of the H3 center. Measuring the integrated absorption coefficient of the zero-phonon line (at 503 nm) of this center at liquid nitrogen temperature suggested a H3 density of 1.7 × 10 18 centers/cm 3 (or 10 ppm). A similarly high concentration of the defect centers was estimated for natural diamond nanocrystallites treated under the same experimental conditions. The centers emitted green light ( λ em = 530 nm) upon illumination by a blue laser ( λ ex = 473 nm or 488 nm). Applications of this novel nanomaterial as fluorescent cellular markers were demonstrated with both confocal fluorescence microscopy and flow cytometry of 70-nm-sized gFND particles uptaken by live HeLa cells through endocytosis.