Detection of Silver and TiO2 Nanoparticles in Cells by Flow Cytometry
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Evaluation of the potential hazard of man-made nanomaterials has been hampered by a limited ability to observe and measure nanoparticles in cells. A FACSCalibur™ flow cytometer and a Stratedigm S-1000 flow cytometer were used to measure changes in light scatter from cells after incubation with either silver nanoparticles (AgNP) or TiO2 nanoparticles. Within the range of between 0.1 μg/mL and 30 μg/mL the nanoparticles caused a proportional increase of the side scatter and decrease of the forward scatter intensity signals. At the lowest concentrations of TiO2 (ranging between 0.1 μg/mL and 0.3 μg/mL), the flow cytometer can detect as few as 5–10 nanoparticles per cell. The influence of nanoparticles on the cell cycle was detected by nonionic detergent lysis of nanoparticle incubated cells that were stained with DAPI or propidium iodide (PI). Viability of nanoparticle treated cells was determined by PI exclusion. Surface plasmonic resonance (SPR) was detected primarily in the far-red fluorescence detection channels after excitation with a 488 nm laser.
Our results suggest that the uptake of nanoparticles within cells can be monitored using flow cytometry. This uptake of nanoparticle data was confirmed by viewing the nanoparticles in the cells using dark-field microscopy. The flow cytometry detection of nanoparticles approach may help fill a critical need to assess the relationship between nanoparticle dose and cellular toxicity. Such experiments using nanoparticles could potentially be performed quickly and easily using the flow cytometer to measure both nanoparticle uptake and cellular health.
Key wordsNanoparticles Flow cytometer Cytometry Toxicology Nanotoxicity Side scatter Plasmonic surface resonance
The manuscript was edited by Enrico Ferrari and Mikhail Soloviev. Thanks are extended to Laura Degn, for her helpful comments and to Wiley & Sons and Springer books for allowing us to reproduce their figures in the publication.
Government Disclaimer: The research described in this chapter has been supported by the US Environmental Protection agency. It has been subjected to agency review and does not necessarily reflect the views of the agency, and no official endorsement should be inferred. The mention of trade names or commercial products does not constitute endorsement or recommendation for use.
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