Thymic stromal lymphopoietin (TSLP) is a cytokine that binds the IL-7-receptor-alpha chain and a unique TSLP receptor (TSLPR) chain. The role of TSLP in human B-cell development has not been elucidated. We show that TSLPR transcripts are expressed most prominently in CD34(+) cells from fetal liver and BM. In general, cell surface expression of TSLPR was low, except on a subset of multilineage-commited progenitor cells. TSLP induced the tyrosine-phosphorylation of STAT5 and the proliferation of multilineage-commited progenitor cells, pro-B cells and pre-B cells. Compared with IL-7, the levels of proliferation after stimulation of the B-cell progenitors with TSLP were lower. Expression of the BCR on the cell surface of fetal cells was inversely correlated to TSLP or IL-7 responsiveness. Pre-B cells from fetal BM, but not fetal liver, were refractory to TSLP or IL-7 stimulation. When employing an in vitro B-cell differentiation culture system starting from CD34(+)CD38(-) multipotent HSC, IL-7 induced a short wave of precursor cell expansion but did not result in long-term survival of mature B cells. TSLP was capable of increasing the proportion and the absolute numbers of more mature human B cells. Overall, we provide evidence that TSLP supports human B-cell differentiation from fetal hematopoietic progenitors.