It has long been unclear how the pluripotent hematopoietic stem cell is restricted to the major lineage progenitors including the progenitors for myeloid, T- and B-cells. This is the result of the absence of a methodology capable of determining the developmental potential of individual progenitors to generate these major lineage cells. We have established such an assay system, termed the multilineage progenitor assay, as a modification of the fetal thymic organ culture system. By examining cells from murine fetal tissues with this assay, we have succeeded in elucidating the process of lineage restrictions in early hematopoiesis.