Endoglin, an ancillary TGF-beta receptor, is differentially expressed in long-term repopulating hematopoietic stem cells (LTR-HSC). Here, we describe simple and highly efficient purification schemes for mouse bone marrow LTR-HSCs using Endoglin as a marker. The Endoglin positive and Sca-1 positive (Endo(Pos) Sca-1(Pos)) population, which contains about 36% of "Side Population" (SP) cells, is highly enriched for LTR-HSCs. In long-term competitive reconstitution assays, 100 such cells reconstituted all lethally irradiated recipients. Interestingly, the Endo(Pos) Sca-1(Pos) population contains comparable LTR-HSC activity in both SP and non-SP fractions, indicating that many HSCs are not captured by the SP phenotype. Furthermore, LTR-HSCs are exclusively found in the Endo(Pos) Sca-1(Pos) Lin(Neg/Low) (lineage negative/low), but not in the Endo(Neg) Sca-1(Pos) Lin(Neg/Low) population, suggesting that the Endo(Pos) population may contain all LTR-HSCs in mouse bone marrow. Finally, we demonstrated that the Endo(Pos) Sca-1(Pos) Rh(Low) (Rhodamine-123 low) phenotype, without using CD34, c-Kit, or Lineage markers, defines a nearly homogenous population of LTR-HSCs.