Quantification of regional cerebral blood flow (CBF) using [O-15]H2O positron emission tomography (PET) requires the use of an arterial input function. Arterial sampling, however, is not always possible, for example in ill-conditioned or paediatric patients. Therefore, it is of interest to explore the use of non-invasive methods for the quantification of CBF. For validation of non-invasive methods, test-retest normal and hypercapnia data from 15 healthy volunteers were used. For each subject, the data consisted of up to five dynamic [O-15]H2O brain PET studies of 10 min and including arterial sampling. A measure of CBF was estimated using several non-invasive methods earlier reported in literature. In addition, various parameters were derived from the time-activity curve (TAC). Performance of these methods was assessed by comparison with full kinetic analysis using correlation and agreement analysis. The analysis was repeated with normalization to the whole brain grey matter value, providing relative CBF distributions. A reliable, absolute quantitative estimate of CBF could not be obtained with the reported non-invasive methods. Relative (normalized) CBF was best estimated using the double integration method.