The Winton Formation provides an important snapshot of Australia's late Mesozoic terrestrial biota, boasting a vertebrate fauna that includes dinosaurs, crocodyliforms, aquatic squamates, turtles, lungfish and teleost fishes, and a flora that has previously been considered to include some of the world's earliest known flowering plants. Despite its significance, poor age control has thus far prevented precise regional and global correlations, limiting the depth of paleobiogeographic assessments. The goal of this study was to use U–Pb isotope dating of detrital zircons by laser ablation to refine the depositional age range of selected horizons within the Winton Formation. We applied this technique, with refined instrumental tuning protocols, to systematically investigate detrital zircon grain ages for five samples from different stratigraphic levels and vertebrate-bearing fossil locations throughout the Winton Formation. Seven different metrics for interpreting the maximum depositional age of each of the detrital zircon samples were compared and our results suggest that sedimentation of the Winton Formation commenced no earlier than latest Albian (~ 103.0–100.5 Ma) and that deposition of the upper vertebrate fossil-rich portion of the section began roughly near or after the Cenomanian–Turonian boundary (93.9 Ma), demonstrating that the formation and its important flora and fauna were deposited primarily during the Late Cretaceous. These results provide a significant advancement in understanding the age of the Winton Formation's flora and fauna, and will help to contextualize Australia's Late Cretaceous terrestrial biota within a broader Gondwanan framework.