To develop non-invasive regional cerebral blood flow (rCBF) measurements using technetium-99m ethyl cysteinate dimer (Tc-99m-ECD) and single-photon emission tomography (SPET), the same graphical analysis as was described in our previous reports using technetium-99m hexamethylpropylene amine oxime (Tc-99m-HMPAO) was applied to time-activity data for the aortic arch and brain hemispheres after intravenous injection of Tc-99m-ECD. Hemispherical brain perfusion indices (BPI) for Tc-99m-ECD showed a highly significant correlation (n = 22, r = 0.935, P = 0.0001) with those for Tc-99m-HMPAO in 11 patients who underwent both tracer studies. Using both linear regression line equations between Tc-99m-ECD BPI and Tc-99m-HMPAO BPI and between Tc-99m-HMPAO BPI and mean cerebral blood flow (CBF) values obtained from a xenon-133 inhalation SPET method in a previous study, Tc-99m-ECD BPI was converted to Xe-133 CBF values (y = 2.60x + 19.8). Then raw SPET images of Tc-99m-ECD were converted to rCBF maps using Lassen's correction algorithm. In this algorithm, the correction factor a was fixed to 1.5, 2.6 and infinite. In the comparison of rCBF values for Tc-99m-ECD SPET with those for Tc-99m-HMPAO SPET in 396 regions of interest in the aforementioned 11 patients, the fixed correction factor a of 2.6 gave nearly the same rCBF values for Tc-99m-ECD (50.1 +/- 16.9 ml/100 g/min, mean + SD) as for Tc-99m-HMPAO (49.9 +/- 17.3 ml/100 g/min). In conclusion, the same non-invasive method as has been used in Tc-99m-HMPAO studies is applicable to a Tc-99m-ECD study for the measurement of rCBF without any blood sampling.