LAUSR

One of the challenges in using in vitro data to understand the potential risks of engineered nanomaterials (ENMs) is that results often differ or are even contradictory among studies. While it is recognized that numerous factors can influence results produced by nanobioassays, there has not yet been a consistently used conceptual framework to identify key sources of variability in these assays. In this paper, we use cause-and-effect analysis to systematically describe sources of variability in four key in vitro nanobioassays: the DCF (2',7'-dichlorofluorescein) assay, an enzyme-linked immunosorbent assay (ELISA) for measuring interleukin-8, a flow cytometry assay (Annexin V/Propidium Iodide), and the Comet assay. These assays measure endpoints that can occur in cells impacted by ENMs through oxidative stress, a principle mechanism for ENM toxicity. The results from this analysis identify control measurements to test for potential artifacts or biases that could occur during conduct of these assays with ENMs. Cause-and-effect analysis also reveal additional measurements that could be performed either in preliminary experiments or each time the assay is run to increase confidence in the assay results and their reproducibility within and among laboratories. The approach used here with these four assays can be used the support the development of a broad range of nanobioassays.