Computed tomography (CT) perfusion is routinely employed to quantitatively assess cerebral hemodynamics and support clinical decision-making in acute ischemic stroke. However, variability in acquisition and reconstruction parameters—particularly slice thickness—remains a significant limitation for the accurate estimation of ischemic core volume. In this retrospective study, the authors investigated the impact of slice thickness on ischemic core quantification by comparing CT perfusion–derived measurements with diffusion-weighted imaging (DWI), used as the reference standard. CT perfusion datasets were reconstructed at slice thicknesses of 1, 3, 5, 7, and 10 mm, and ischemic core volumes were automatically calculated using thresholds of relative cerebral blood flow (rCBF) and relative cerebral blood volume (rCBV). Quantitative agreement with DWI was assessed through mean absolute error (MAE), Bland–Altman analysis, and intraclass correlation coefficient (ICC). Among the 87 patients included, the lowest estimation error was achieved using an rCBF threshold of <38%. Reconstructions with slice thicknesses of 3, 5, and 7 mm showed the highest concordance with DWI, showing minimal bias and strong reproducibility. In contrast, both very thin (1 mm) and thicker (10 mm) reconstructions were associated with reduced accuracy. Overall, these findings suggest that CT perfusion scans reconstructed with intermediate slice thicknesses (3–7 mm) provide the most reliable estimation of ischemic core volume, supporting a more standardized approach to image reconstruction in the acute stroke setting.