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Background: It remains a great challenge to measure systemic blood flow in critically ill newborns. In a former study we validated the modified carbon dioxide Fick (mCO₂F) method for measurement of cardiac output in a newborn lamb model. In this new study we studied the influence of a left-to-right shunt on the accuracy of the mCO₂F method. Objective: To analyze the influence of a left-to-right shunt on the agreement between cardiac output measurement with the mCO₂F method and ultrasonic transit time pulmonary blood flow in a lamb model. Methods: The study was approved by the Ethical Committee on Animal Research of the Radboud University Nijmegen and performed in 8 random-bred lambs. A Gore-Tex® shunt was placed between the left pulmonary artery and the descending aorta. This aortopulmonary shunt was intermittently opened and closed, while cardiac output was manipulated by creating hemorrhagic hypotension. Cardiac output measurement with the mCO₂F method (Q^mCO2F) was compared with pulmonary blood flow obtained by a transit time ultrasonic flow probe positioned around the common pulmonary artery (Q^APC). Results: Bias, defined as Q^mCO2F – Q^APC, was calculated for each measurement. With an open shunt there was a significant left-to-right shunt (mean Qp/Qs ratio 2.26; range 1.56–3.69). Mean bias (SD) was –12.3 (50.4) ml·kg^–1·min^–1 and –12.3 (42.7) ml·kg^–1·min^–1 for measurements with a closed and open shunt, respectively (no statistical significant difference). Conclusions: Cardiac output measurement with the mCO₂F method is reliable and easily applicable in ventilated newborn lambs, also in the presence of a significant left-to-right shunt.