Interaction between Fluorescent Nanodiamond and Human Transferrin and Intracellular Imaging

Abstract

Nanodiamond (ND), as a member of carbon nanomaterials family, has recently received increasing attention for their potential applications as imaging and drug delivery agents. Due to several charming properties, such as surface functionalization capability, biocompatibility and chemical stability, ND demonstrates high affinity to biomolecules. In this work, the adsorptive behavior of oxidized fluorescent nanodiamond (FND, with a size of ca. 140 nm) for hTf was investigated. The amount of surface carboxylic acid on oxidized FND was 126 μmol/g determined by conductometric backward titration. This amount is 29.7% of the total surface atoms. HTf physically adsorbed on the surface of FND (FND-hTf) in PBS (pH7.4) shows that the isothermyal adsorption behavior is coincident with Langmuir model and the maximum adsorbed amount is (176.46±2.13) μg/mg. ND coated human transferrin (hTf) can improve the dispersity and stability compared to pristine FND under a physiological environment or in cell culture medium observed through optical microscope and be more suitable for biomedicine applications. Simultaneously, pH effect on hTf adsorbed onto FND is also inquiried. The result exhibits that FND has the greatest adsorption capacity for hTf close to the isoelectric point. Owing to the negatively charged nitrogenvacancy (N-V)defect centers, FND can absorb strongly at ca. 560 nm and emit fluorescence efficiently at ca. 700 nm, which can be well quantitatively and qualitatively analyzed by flow cytometry and confocal fluorescence images. In vitro experiments of human liver cancer cells (HepG2) uptake of nanoparticles display that FND-hTf nanoparticles are more easily endocytosed than that of pristine FND. So FND-hTf is conducive to cell imaging. Furthermore, flow cytometry assay indicates cellular uptake of FND-hTf reached a plateau at about 8 h and the uptake half-life is about (1.41±0.22) h at a particle concentration of 100 μg/mL. The results obtained by confocal fluorescent images display that the FND-hTf nanoparticles locate in the cytoplasm and mainly distribute on the surface of the nucleus, however, can not enter the nucleus.