Mechanisms that govern hematopoietic lineage specification, as opposed to the expansion of committed hematopoietic progenitors, from human pluripotent stem cells (hPSCs) have yet to be fully defined. Here, we show that within the family of genes called inhibitors of differentiation (ID), ID1 and ID3 negatively regulate the transition from lineage-specified hemogenic cells to committed hematopoietic progenitors during hematopoiesis of both human embryonic stem cells (hESCs) and human induced pluripotent stem cell (hiPSCs). Upon hematopoietic induction of hPSCs, levels of ID1 and ID3 transcripts rapidly increase, peaking at the stage of hemogenic precursor emergence, and then exclusively decrease during subsequent hematopoietic commitment. Suppression of ID1 and ID3 expression in hemogenic precursors using specific small interfering RNAs augments differentiation into committed hematopoietic progenitors, with dual suppression of ID1 and ID3 further increasing hematopoietic induction compared with upon knockdown of each gene alone. This inhibitory role of ID1 and ID3 directly affects hemogenic precursors and is not dependent on non-hemogenic cells of other lineages within developing human embryoid bodies from hESCs or hiPSCs. Our study uniquely identifies ID1 and ID3 as negative regulators of the hPSC-hematopoietic transition from a hemogenic to a committed hematopoietic fate, and demonstrates that this is conserved between hESCs and hiPSCs.