A common technique for calculating cerebral blood flow (CBF) and mean transit time (MTT) is to track a bolus of contrast agent using perfusion-weighted MRI (PWI) and to deconvolve the change in concentration with an arterial input function (AIF) using singular value decomposition (SVD). This method has been shown to often overestimate the volume of tissue that infarcts and in cases of severe vasculopathy to produce CBF maps that are inconsistent with clinical presentation. This study examines the effects of tracer arrival time differences between tissue and a user-selected global AIF on flow estimates. CBF and MTT were calculated in both numerically simulated and clinically acquired PWI data where the AIF and tissue signals were shifted backward and forward in time with respect to one another. Results show that when the AIF leads the tissue, CBF is underestimated independent of extent of delay, but dependent on MTT. When the AIF lags the tissue, flow may be over- or underestimated depending on MTT and extent of timing differences. These conditions may occur in practice due to the application of a user-selected AIF that is not the “true AIF” and therefore caution must be taken in interpreting CBF and MTT estimates. Magn Reson Med 50:856–864, 2003. © 2003 Wiley-Liss, Inc.