ABSTRACT
Fluorescent nanodiamond (FND) contains nitrogen-vacancy defect centers as fluorophores. The intensity of its fluorescence can be significantly enhanced after deposition of the particle (35 or 140 nm in size) on a nanocrystalline Ag film without a buffer layer. The excellent photostability (i.e. neither photobleaching nor photoblinking) of the material is preserved even on the Ag film. Concurrent decrease of excited state lifetimes and increase of fluorescence intensities indicate that the enhancement results from surface plasmon resonance. Such a fluorescence enhancement effect is diminished when the individual FND particle is wrapped around by DNA molecules, as a result of an increase in the distance between the color-center emitters inside the FND and the nearby Ag nanoparticles. A fluorescence intensity enhancement up to 10-fold is observed for 35 nm FNDs, confirmed by fluorescence lifetime imaging microscopy.
ABSTRACT
Fluorescent nanodiamond is a new nanomaterial that possesses several useful properties, including good biocompatibility, excellent photostability and facile surface functionalizability. Moreover, when excited by a laser, defect centres within the nanodiamond emit photons that are capable of penetrating tissue, making them well suited for biological imaging applications. Here, we show that bright fluorescent nanodiamonds can be produced in large quantities by irradiating synthetic diamond nanocrystallites with helium ions. The fluorescence is sufficiently bright and stable to allow three-dimensional tracking of a single particle within the cell by means of either one- or two-photon-excited fluorescence microscopy. The excellent photophysical characteristics are maintained for particles as small as 25 nm, suggesting that fluorescent nanodiamond is an ideal probe for long-term tracking and imaging in vivo, with good temporal and spatial resolution.
Subject(s)
Contrast Media/chemistry , Crystallization/methods , Diamond/chemistry , Image Enhancement/methods , Microscopy, Fluorescence/methods , Nanoparticles/chemistry , HeLa Cells , Humans , Materials TestingABSTRACT
Type Ib diamonds emit bright fluorescence at 550-800 nm from nitrogen-vacancy point defects, (N-V)(0) and (N-V)(-), produced by high-energy ion beam irradiation and subsequent thermal annealing. The emission, together with noncytotoxicity and easiness of surface functionalization, makes nano-sized diamonds a promising fluorescent probe for single-particle tracking in heterogeneous environments. We present the result of our characterization and application of single fluorescent nanodiamonds as cellular biomarkers. We found that, under the same excitation conditions, the fluorescence of a single 35-nm diamond is significantly brighter than that of a single dye molecule such as Alexa Fluor 546. The latter photobleached in the range of 10 s at a laser power density of 10(4) W/cm(2), whereas the nanodiamond particle showed no sign of photobleaching even after 5 min of continuous excitation. Furthermore, no fluorescence blinking was detected within a time resolution of 1 ms. The photophysical properties of the particles do not deteriorate even after surface functionalization with carboxyl groups, which form covalent bonding with polyL-lysines that interact with DNA molecules through electrostatic forces. The feasibility of using surface-functionalized fluorescent nanodiamonds as single-particle biomarkers is demonstrated with both fixed and live HeLa cells.
