OBJECTIVE: We sought to evaluate the frequency of CD34⁺ hematopoietic progenitor/stem cells across gestation and the proliferative response of early versus late gestational age fetal blood to growth factor stimulation. STUDY DESIGN: Fetal blood samples were obtained at 17 to 41 weeks' gestational age. The mononuclear cell population was separated by red blood cell lysis. The frequency of CD34⁺ cells within the lymphocyte-monocyte light scatter gate and the expression of CD34 by colony-forming cells of different types were determined by fluorescence-activated cell sorting. The growth response of colony-forming cells to varying concentrations of defined growth factors (interleukin-3, interleukin-6, granulocyte-macrophage colony-stimulating factor, stem cell factor, and erythropoietin) was determined, as well as the frequency of burst-forming units erythroid, colony-forming units granulocyte-macrophage, and total colony-forming cells. Samples from 17 to 24 weeks' gestation (early) were compared with those of 39 to 41 weeks' gestation (late). RESULTS: The frequency of CD34⁺ cells in early fetal blood (17 to 24 weeks' gestation) was 4.9-fold higher (6.4% vs 1.3%, p < 0.002) than term gestation (37 to 41 weeks' gestation) and declined linearly with gestational age (p < 0.0001). When gestational ages were grouped into 4-week blocks (17 to 20, 21 to 24, 25 to 28, 29 to 32, 33 to 36, and >37 weeks), statistically significant changes in the frequency of CD34⁺ cells did not occur until after 28 weeks' gestation. By cell sorting, >99% of all colony-forming cells were contained within the CD34⁺ population at all gestational ages tested. Compared with term, the frequency of colony-forming cells was significantly greater in early fetal blood (burst-forming units erythroid [18.1-fold, p < 0.0001], colony-forming units granulocyte-macrophage [2.9-fold, p <0.001], and total colony-forming cells [9.4-fold, p < 0.0001]). However, when the frequency of colony-forming cells was corrected for the frequency of CD34⁺ cells, only the number of burst-forming units erythroid remained significantly greater in early fetal blood. The relative size of the colonies formed by individual progenitors from early fetal blood was greater than those from term samples. CONCLUSIONS: From 17 to 41 weeks' gestation the frequency of CD34⁺ cells in fetal blood, which includes hematopoietic progenitors and stem cells, decreases. This decline occurs during the transition from hepatic to bone marrow hematopoiesis. Early fetal blood, with a higher circulating frequency of progenitor/stem cells and proliferative capacity, may be a preferable target for gene therapy. (Am J Obstet Gynecol 1998;178:931-7.)