Maternal serum screening to identify fetal aneuploidies is now routinely offered during the second trimester of pregnancy in developed countries. The purpose of this prospective study was to assess the value of maternal serum screening between 15 and 20 weeks of gestation to detect fetal aneuploidies and to determine the false positive rate (FPR). Blood samples were collected from 1,062 pregnant women between 15 and 20 weeks of gestation. Samples were assayed for alpha-fetoprotein (AFP), free beta human chorionic gonadotropin (beta-hCG) and unconjugated estriol (uE3). Medians were established at each week from 200 normal, singleton pregnancies. Second trimester risk was calculated using the maternal age and different combinations of AFP, beta-hCG and uE3. Screening results calculated by likelihood ratio to be equal to or greater than 1:270 were considered positive. If the gestational age was confirmed by ultrasonography, genetic counselling and amniocentesis were offered. Ten fetal chromosomal abnormalities were detected with maternal serum screening. Sample's size does not allow a correct detection rate estimation, but false positive rate (FPR) was found to be 6.5%. This FPR has a clinical application. At a cut-off of 1:270, second trimester screening best results were obtained using a combination of all three biochemical markers. These results confirm the efficacy of maternal serum screening for fetal chromosomal abnormalities with a low FPR. The measurement of AFP, beta-hCG and uE3 is an effective prenatal screening test.