This cruise report is a summary of a field survey conducted within two Great Lakes Region Areas of Concern (AOCs), Ashtabula River and Milwaukee Estuary, August 18 – 25, 2012. Synoptic sampling of multiple ecological indicators was conducted in the benthos and the water column at each of 30 stations throughout these two AOCs using a random probabilistic sampling design. Twelve of the 30 stations were located in the Ashtabula River, Ohio and the remaining 18 were located in the Milwaukee Estuary and its tributaries. At each station samples were collected for the analysis of benthic macroinfauna community structure and composition; concentrations of chemical contaminants in sediments; sediment toxicity; and other basic habitat characteristics such as temperature, dissolved oxygen, turbidity, pH, sediment grain size, and organic carbon content. Other indicators, from a human-dimension perspective, were also recorded, including presence of surface trash, visible oil sheens in sediments or water, or noxious/oily sediment odors.