An electron microscopy based method for the detection and quantification of nanomaterial number concentration in...

Improved detection and characterization of nanomaterials (NMs) in complex environmental media requires the development of novel sampling approaches to improve the detection limit to be close to environmentally realistic concentrations. Transmission electron microscopy (TEM) is an indispensable metrological tool in nanotechnology and environmental nanoscience due to its high spatial resolution and analytical capabilities when coupled to spectroscopic techniques. However, these capabilities are hampered by the conventional sample preparation methods, which suffer from low NM recovery. The current work presents a validated, fully quantitative sampling technique for TEM that overcomes conventional sample preparation shortcomings, and thus enables the use of TEM for measurement of particle number concentration and their detection in complex media at environmentally realistic concentrations. This sampling method is based on ultracentrifugation of NMs from suspension onto a poly-l-lysine (PLL) functionalized TEM grid, using active deposition (by ultracentrifugation) and retention (by PLL interactions with NM surface) of NMs on the substrate, enabling fully quantitative analysis. Similar analysis with AFM was satisfactory in simple media but the lack of chemical-selectivity of AFM limits its applicability for the detection of NMs in complex environmental samples. The sampling approach was validated using both citrate- and PVP-coated AuNMs in pure water, which demonstrated an even distribution of NM on the TEM grid and high NM recovery (80-100%) at environmentally relevant NM concentrations (ca. 0.20-100 μg L(-1)). The applicability of the sampling method to complex environmental samples was demonstrated by the quantification of particle number concentration of AuNMs in EPA soft water (with and without Suwannee River fulvic acid) and lake water. This sample preparation approach is also applicable to other types of NMs with some modifications (e.g. centrifugation force and time) to insure full sample recovery. This TEM sampling method is key to the accurate quantification of NM number concentration, and therefore to improving our understanding of environmental fate, behavior, effects and dose of NMs.