An enzyme-linked immunosorbent assay (ELISA), used to detect IgG and IgM antibody specific for human parvovirus B19, was established by use of human parvovirus B19 capsid protein VP-1 expressed in Escherichia coli. Paired samples of serum derived from 44 mothers and single samples derived from 24 babies having unexplained non-immune hydrops fetalis (NIHF), were tested by means of the assay. Five cases (11%) of NIHF were suspected of having been induced by intrauterine human parvovirus B19 infection because the samples of maternal serum were positive for parvovirus B19 IgM antibody. Four of the five cases arose during an outbreak of erythema infectiosum. According to our study and previous reports over 90% of NIHF caused by parvovirus B19 intrauterine infection have become clinically overt in the second trimester of pregnancy during a period ranging from 15 to 27 weeks of gestation.