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Preparation, characterization, and in vitro dosimetry of dispersed, engineered nanomaterials

Nature Protocols volume 12, pages 355–371 (2017)Cite this article

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Abstract

Evidence continues to grow of the importance of in vitro and in vivo dosimetry in the hazard assessment and ranking of engineered nanomaterials (ENMs). Accurate dose metrics are particularly important for in vitro cellular screening to assess the potential health risks or bioactivity of ENMs. To ensure meaningful and reproducible quantification of in vitro dose, with consistent measurement and reporting between laboratories, it is necessary to adopt standardized and integrated methodologies for (i) generation of stable ENM suspensions in cell culture media; (ii) colloidal characterization of suspended ENMs, particularly of properties that determine particle kinetics in an in vitro system (size distribution and formed agglomerate effective density); and (iii) robust numerical fate and transport modeling for accurate determination of the ENM dose delivered to cells over the course of the in vitro exposure. Here we present an integrated comprehensive protocol based on such a methodology for in vitro dosimetry, including detailed standardized procedures for each of these three critical aims. The entire protocol requires ∼6–12 h to complete.

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Figure 1: Harvard dispersion dosimetry protocol (HDDP).
Figure 2: ENM dispersion protocol.
Figure 3: Volumetric centrifugation method (VCM).
Figure 4: Determination of DSEcr value.
Figure 5: Volume size distribution.
Figure 6: Fate and Transport modeling results.

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Acknowledgements

This research project was supported by a Harvard–National Institute of Environmental Health Sciences Nanosafety Center grant (1U24ES026946) and a National Science Foundation grant (1436450), both awarded to P.D.

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  1. Department of Environmental Health, Center for Nanotechnology and Nanotoxicology, HSPH-NIEHS Nanosafety Research Center, Harvard School of Public Health, Boston, Massachusetts, USA

    Glen M DeLoid, Joel M Cohen, Georgios Pyrgiotakis & Philip Demokritou

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  1. Glen M DeLoid
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G.M.D., J.M.C., G.P., and P.D. co-wrote the manuscript. P.D. oversaw the manuscript preparation.

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Correspondence to Philip Demokritou.

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Supplementary information

Supplementary Software

Supplementary Software. This file contains the MATLAB .m code to run the distorted grid computational dosimetry model as described in the text. (TXT 40 kb)

Supplementary Data

Supplementary Data. This file contains the dosimetry output obtained from running the distorted grid computational model with parameters set as described in the text for an example Fe2O3 ENM. (XLSX 1920 kb)

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DeLoid, G., Cohen, J., Pyrgiotakis, G. et al. Preparation, characterization, and in vitro dosimetry of dispersed, engineered nanomaterials. Nat Protoc 12, 355–371 (2017). https://doi.org/10.1038/nprot.2016.172

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