In this study we report a new triple test that combines serum AFP, urine β€-core fragment of hCG, total urine estriol, and maternal age for calculating individual Down syndrome odds in the second trimester. The urine β€-core fragment/estriol ratio was used as a single screening variable. Analyte levels were measured prospectively in 10 Down syndrome cases and 346 normals. Individual Down syndrome odds were calculated by multiplying the product of the Down syndrome likelihood ratios of serum AFP and urine β€-core/estriol levels by the age-related midtrimester risk. The screening efficiency of an algorithm that combines urine β€-core/estriol with maternal age was compared to one that included serum AFP data. A 90% detection rate for Down syndrome was obtained at a 4.65% false positive rate. This was superior to the 75% sensitivity at 5% false positive rate observed when β€-core/estriol and age alone were used. This new triple test has a higher screening efficiency than that generally reported for the traditional serum triple screen and other urine tests, and it also provides information on the risk of neural tube defects. If confirmed in larger trials, the new algorithm could be used as an alternative to the traditional serum triple screen.