Cytocompatibility and 3D biodistribution with oxidized nanographene assessed by digital holographic microscopy

Nano graphene-based materials offer interesting physicochemical and biological properties for biotechnological applications due to their small size, large surface area and ability to interact with cells/tissues. Among carbon-based nanomaterials, graphene oxide is one of the most used in biological field. There is an increasing interest in shedding light on the interaction mechanisms of nanographene oxide (nGO) with cells. In fact, the effects on human health of GO, and its toxicological profile, are still largely unknown. Here we show that, by minimizing the oxidation degree of GO, its toxicity is significantly reduced in NIH 3T3 cells. Moreover, we show that mild oxidation of graphene nanoplatelets produces nGO particles, which are massively internalized into the cell cytoplasm. MTT(3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide) assay was performed to analyze cell viability. Transmission electron microscopy (TEM) analysis was performed to evaluate nGO internalization mechanism into the cytoplasm under different oxidation degree and concentrations. For the first time, we evaluated quantitatively, the cell volume variation after nGO internalization in live fibroblasts through a label-free digital holography (DH) imaging technique and in quasi-real-time modality, thus avoiding the time-consuming and detrimental procedures usually employed by electron-based microscopy. In conclusion, here we have demonstrated that DH can be a viable tool to visualize and display 3D distributions of nano graphene oxide (nGO) uptake by fibroblast cells. DH opens the route for high-throughput investigation at single cell level for understanding how in different conditions nanoparticles aggregates distribute inside the cells.