Objectives: To construct reference intervals for fetal lung volumes measured longitudinally using three-dimensional (3D) ultrasound, and to evaluate the effect of gender on lung size. Methods: This was a prospective, longitudinal study in the obstetric outpatient department of the VU University Medical Center, Amsterdam. Seventy-eight women with uncomplicated pregnancies were scanned three to four times at gestational ages of 18-34 weeks. 3D models of the lung were constructed using the ultrasound machine's software. After the infants were delivered the entire group was reanalyzed with regard to fetal gender. Centiles for the lung volumes of the entire group and for each gender separately were estimated using multilevel modeling. Results: Charts and tables of right and left fetal lung volumes, using gestational age and estimated fetal weight as the independent variables, are presented. There was a significant difference in lung volume between male and female fetuses at each gestational age. Charts and tables of right and left fetal lung volumes for each gender at gestational ages of 18-34 weeks are also presented. Conclusions: We present valid references for volumetric measurements of the right and left fetal lungs in male and female fetuses. The feasibility and reliability of fetal lung volume measurements using 3D ultrasound is good. Copyright © 2006 ISUOG.