The aims of the studies reviewed were 1) to identify the molecular forms of acetylcholinesterase encountered in amniotic fluid from pregnancies with a normal fetus and those with a fetal open neural tube defect or other fetal malformation and, 2) to raise and characterize antibodies against human acetylcholinesterase and to identify those useful for immunochemical determination of amniotic fluid acetylcholinesterase where there is a fetal open neural tube defect. Eleven monoclonal antibodies and one polyclonal rabbit antibody were evaluated with regard to their clinical usefulness in the antenatal diagnosis of open neural tube defects. One of these, the monoclonal antibody 4F19, preferentially bound acetylcholinesterase from human brain and identified better than the others amniotic fluid samples from pregnancies with a fetal open neural tube defect (I, II). The monoclonal antibody 4F19 was used in an enzyme antigen immunoassay whose performance was found to be similar to that of the polyacrylamide electrophoretic gel test for acetylcholinesterase determination (III). However, the 4F19 enzyme antigen immunoassay is simpler, more rapid and less technically demanding than the gel test, and furthermore, it gives a quantitative result. The 4F19 enzyme antigen immunoassay was also compared with the alpha-fetoprotein test, normally used as the primary test for the antenatal diagnosis of open neural tube defects. The 4F19 enzyme antigen immunoassay performed better than the alpha-fetoprotein test, but the best performance was found for a combination of the two tests (VI). A positive result can be found using the combined tests for conditions other than open neural tube defects, e.g. abdominal wall defects, intrauterine fetal death and other fetal malformations. These conditions can often be discerned by ultrasound examination. However, combining the result of the 4F19 enzyme antigen immunoassay with the result of an enzyme antigen immunoassay for butyrylcholinesterase makes a discrimination between these conditions possible (V). The diagnostic implications of the above procedures are evaluated and specific recommendations concerning their use are given.