In the first half of pregnancy, patterns of esterase activity, as well as electrophoretic esterase isozyme profiles of trophoblast (fetal placenta) homogenates can be distinguished from those of other embryonic tissues and from those of the decidua (maternal placenta). Two electrophoretic bands of esterase activity are found in trophoblast but not embryo proper; one is of maternal origin. Both bands are found in ectopic pregnancies containing mainly giant cells, and at least one is present in preimplantation late morulae and blastocysts. Blastocysts cultured in modified Eagle medium containing dialysed serum give rise mainly to trophoblast-like growths, and under these conditions, the esterase profile is predominantly trophoblast-like. Embryos cultured from the two-cell stage or viable blastocysts which have been unable to hatch from their zona pellucidae also produce trophoblast-like esterase. Unmodified Eagle medium supplemented with heat-inactivated serum allows the proliferation of cells that appear to derive from the inner cell mass. Under these conditions, the esterase profiles closely resemble those of the yolk sac and embryo proper, even after 3 weeks in culture.