North America's Atlantic Coast has been a focus of human settlement and subsistence for millennia, but sea-level rise, sedimentation, and other processes pose significant challenges for archaeological research. Radiocarbon dating of 31 shell middens near the Rhode River Estuary, Maryland provides an opportunity to evaluate human land use, settlement, and cultural chronologies on the Chesapeake Bay. Sixty calibrated radiocarbon dates on eastern oyster (Crassostrea virginica) shell and charcoal demonstrate that Native Americans, colonial, and historic peoples harvested oysters and other shellfish from at least 3200 years ago through the 19th century. The number of dated sites increases during the Late Woodland period after about 1000 cal yr B.P., a factor probably related to greater site visibility and preservation, as well as increased human exploitation of the watershed. Accumulation rates for five of the shell middens provide preliminary indications that some of the sites accumulated rapidly suggesting, along with other evidence, that many of the region's shell middens were logistical or perhaps seasonal camps. Our study demonstrates the importance of regional watershed surveys and radiocarbon dating programs to help build and refine cultural chronologies in coastal regions threatened by sea-level rise and other processes.