Objective: The objective of this study is to evaluate whether assessment pulmonary vascular reactivity in response to maternal hyperoxygenation using fractional moving blood volume (FMBV) is associated with lesser variability between individual measurements than what is observed with direct Doppler measurements. Study design: Forty-five measurements were performed in 15 singleton fetuses with normal lung development at three time points in the latter half of pregnancy (range: 25.9-36.7weeks). We further evaluated five fetuses with severe congenital diaphragmatic hernia. Lung perfusion was assessed using power Doppler ultrasound, and images were stored for offline FMBV calculation, both at base line and during oxygen administration (9L/min for 10min). The proportionate difference between both measurements is further referred to as deltaFMBV. Results: Overall, 91% of images were of sufficient quality for further analysis. There was no correlation between pulmonary reactivity to oxygen (deltaFMBV) and gestational age in controls (12.9±32.1%). Moreover, deltaFMBV showed large variability between subjects, as well as within the same fetus throughout gestation. We observed good intraobserver (0.88; 0.84) and interobserver (0.88; 0.77) reproducibility for both controls and congenital diaphragmatic hernia, respectively (intraclass correlation coefficients). Conclusion: Despite being a reproducible method to study the lung vasculature, the large variability of FMBV following hyperoxygenation limits its clinical translation.