Coronary flow reserve (CFR) is an important index of coronary microcirculatory function. The objective of this study was to validate the reproducibility and accuracy of intravascular conductance catheter-based method for measurements of baseline and hyperemic coronary flow velocity (and hence CFR). The absolute coronary blood velocity was determined by measuring the time of transit of a saline injection between two pairs of electrodes (known distance) on a conductance catheter during a routine saline injection without the need for reference flow. In vitro validation was made in the velocity range of 5 to 70 cm/s in reference to the volume collection method. In 10 swine, velocity measurements were compared with those from a flow probe in coronary arteries at different CFR attained by microsphere embolization. In vitro, the mean difference between the proposed method and volume collection was 0.7 Ϯ 1.34 cm/s for steady flow and Ϫ0.77 Ϯ 2.22 cm/s for pulsatile flow. The mean difference between duplicate measurements was 0 Ϯ 1.4 cm/s. In in vivo experiments, the flow (product of velocity and lumen cross-sectional area that is also measured by the conductance catheter) was determined in both normal and stenotic vessels and the mean difference between the proposed method and flow probe was Ϫ1 Ϯ 12 ml/min (flow ranged from 10 to 130 ml/min). For CFR, the mean difference between the two methods was 0.06 Ϯ 0.28 (range of 1 to 3). Our results demonstrate the reproducibility and accuracy of velocity and CFR measurements with a conductance catheter by use of a standard saline injection. The ability of the combined measurement of coronary lumen area (as previously validated) and current velocity and CFR measurements provides an integrative diagnostic tool for interventional cardiology.catheter; conductance ISCHEMIC HEART DISEASE is complex and often involves changes in epicardial coronary arteries and myocardial microvasculature. Coronary flow reserve (CFR, ratio of maximal to baseline flow) accounts for both epicardial and microvascular resistances (7). In the absence of epicardial artery disease, the CFR provides an index of the health of coronary microcirculation. Many patients who undergo percutaneous coronary interventions and revascularizations have low coronary flow because of both epicardial and microcirculatory disease (15,28).Several methods have been used for measuring velocity or blood flow rate in animals and humans. The intracoronary Doppler wire is a well-established method for determining blood flow velocity (3,11,19). The measurement relies on the detection of red blood cells moving past the ultrasound emitter/ receiver at the end of a guide wire. Although this method has been used clinically, it is angle dependent, and especially in disease vessels, it lacks the accuracy and reproducibility needed for a reliable diagnosis (17). The thermodilution method has also been proposed for determining blood velocity (24), but it has not garnered significant clinical utility because it requires a reference injection f...