We assessed the qualitative test for AChE in addition to AFP for the diagnosis of NTD in 420 AF samples selected from 16,000 second trimester amniocenteses. Three-hundred and thirty were from normal and ninety from abnormal pregnancies. There were three false positives by this test, two from stored samples containing fetal blood and one from a clear specimen; all had normal outcomes and two had had abnormal AFPs. Forty-seven of forty-seven NTDs were positive by AChE and included 26 cases of anencephaly, 16OSBs, and 5 encephaloceles. In addition, about half of the cases of omphalocele, fetal demise, cystic hygroma, and hydrops gave positive results. One case of congenital nephrosis was negative. We conclude that AChE gel electrophoresis is a useful adjunctive test when AFP levels measure +3 SD above the mean. Although no false negative occurred in this series identified by elevated AFP or increased risk for NTDs, the number in unselected pregnancies is not known.