Computed tomography (CT) perfusion can analyze tissue perfusion and quantitative parameters, including blood flow, blood volume, and transit time. CT perfusion has been used for evaluating split renal function. However, its applicability in veterinary medicine was not validated. This study aimed to evaluate the correlation of renal blood flow (RBF) derived by maximum slope-based CT perfusion and an ultrasonic flow probe and assess the effect of the presence of a pre-existing contrast medium on CT perfusion in the kidneys. In five healthy purpose-bred beagles, CT perfusion was performed at the level of the left renal hila after injection of 1 mg/kg iohexol, during measuring RBF with an ultrasonic flow probe placed on the left renal artery. After post-contrast CT scan with injection of 2 mg/kg iohexol, CT perfusion scan was repeated with the same protocol used in the first perfusion study. The CT perfusion derived RBF was analyzed based on the maximum slope and was compared with the true RBF obtained using an ultrasonic flow probe. Results indicated that CT perfusion derived RBF was significantly correlated with true RBF, although CT perfusion derived RBF did not match the absolute value of the true RBF. It was correlated with the true RBF, even in the presence of a pre-existing contrast medium in the kidney. CT perfusion can estimate the change in individual renal perfusion non-invasively, and this method can be used supplementary to the conventional CT protocol in clinic.