Abstract Background Thymic stromal derived lymphopoietin (TSLP) is preferentially and highly expressed in the thymus, but its function in T cell development is not clear. Results We report here that TSLP, independently or in combination with IL-7, enhances thymopoiesis in the murine fetal thymic organ culture (FTOC) model. Furthermore, TSLP preferentially increases the number and proliferation of the (DN1 and DN2) pro-T progenitor cells, and FTOC lobes from TSLP receptor-null mice show a decreased number of these cells. Finally, DN1-DN2 cells expanded with TSLP in vitro are functional T progenitors that are able to differentiate into mature T cells in fetal or adult thymus organs. Conclusion Together, these data suggest that TSLP plays an important role in expansion of thymocyte progenitors and may be of value for expanding T progenitor cells in vitro.