To elucidate the role of helix-loop-helix (HLH) Id proteins in hematopoietic differentiation, we used a model of embryonic stem (ES) cell differentiation in vitro which gives access not only to hematopoietic myeloid progenitor cells but also to the more primitive blast colony-forming cell (BL-CFC), the in vitro equivalent of the hemangioblast that gives rise to blast cell colonies in the presence of VEGF. We first demonstrated that ES cell-derived blast cell colonies could be used as a model to study hematopoietic differentiation and maturation. We next established the expression profile of Id genes in this model. Transcripts of the four Id genes were present in ES cells. Id1, Id3 and Id4 gene expression was down-regulated during the development of blast cell colonies while that of Id2 was maintained. Thus, Id1, Id3, and Id4 proteins are candidates for being negative regulators of hematopoiesis in the model of hematopoietic ES cell differentiation in vitro.