Hematopoietic stem cells (HSCs) are thought to divide infrequently based on their resistance to cytotoxic injury targeted at rapidly cycling cells^, and have been presumed to retain labels such as the thymidine analog 5-bromodeoxyuridine (BrdU). However, BrdU retention is neither a sensitive nor specific marker for HSCs. Here we show that transient, transgenic expression of a histone 2B (H2B)–green fluorescent protein (GFP) fusion protein in mice has several advantages for label-retention studies over BrdU, including rapid induction of H2B-GFP in virtually all HSCs, higher labeling intensity and the ability to prospectively study label-retaining cells, which together permit a more precise analysis of division history. Mathematical modeling of H2B-GFP dilution in HSCs, identified with a stringent marker combination (L⁻K⁺S⁺CD48⁻CD150⁺), revealed unexpected heterogeneity in their proliferation rates and showed that ∼20% of HSCs divide at an extremely low rate (≤0.8–1.8% per day).