We recently showed that long-term marrow cultures can be used to demonstrate the presence of Philadelphia (Ph1) negative progenitors in patients with newly diagnosed Ph1-positive chronic myeloid leukemia (CML). We now report results for 6 chronic phase patients studied 5–83 mo postdiagnosis and an additional 3 newly diagnosed patients. Marrow metaphases were exclusively Ph1-positive. Clonogenic assays revealed a minor population of Ph1-negative progenitors in 3 cases (1 treated, 2 untreated). Long-term marrow culture adherent layers contained Ph1- negative progenitors in 6 cases (3 treated, 3 untreated). Whenever this occurred, the Ph1-negative population had become the only one detectable within 3–4 wk, and this was always associated with a rapid decline of the Ph1-positive population. For 2 of the 3 cases where Ph1- negative progenitors were not detected, there was a similar rapid decline in the Ph1-positive population in culture. In the other case, Ph1-positive progenitors were maintained at levels typically seen in normal long-term marrow cultures. These results suggest that chromosomally normal stem cells may persist for a considerable period in the marrow of some, but perhaps not all, patients with CML, even in the face of maintenance chemotherapy. In addition, they provide new evidence of heterogeneity in this disease, as shown by the variable ability of Ph1-positive progenitor populations to be maintained in vitro.