Chemical contaminants are introduced to environmental waters via many sources, and many of these contaminants have the potential to adversely affect organisms living in these waters. Recognized adverse effects include the induction of cancer via genotoxic mechanisms, endocrine disruption via the derailment of normal hormone signaling pathways, and outright toxicity leading to disease or death. Scientists in the U.S. Geological Survey (USGS) within DOI have established or adapted water collection, extraction, and in vitro screening assays to evaluate the bioactivity of surface water samples. These approaches circumvent the need to utilize vertebrates and minimize endpoint variability in bioactivity measures. Data from these assays are incorporated into predictive modeling analyses to identify land uses associated with predicted biological disruption. In addition, they are also applied to responsibly inform site selection for comprehensive environmental sampling. During 2018-2019 these assays were applied to augment USGS and other collaborator data sets collected from environmental surface and well waters collected in the eastern and midwestern United States.