Quantitation of Hoechst 33342 SP cells in bone marrow expansion cultures after MDR1 gene transfer.

Murine bone marrow cells were prestimulated for 48 hours in the presence of growth factors and then retrovirally transduced with either the HaMDR1 or the HaDHFR^L22Y vector for another 48 hours. The time point immediately after coculture was defined as day 0 of expansion. Transduced cells were then grown in suspension cultures for 12 days. Hoechst 33342 staining of BM cells was performed on days 0, 6, and 12 to determine the frequency of SP cells within the expanding cell populations. FACS profiles, representative of 3 independent expansion experiments, are shown for BM cells transduced with either the HaMDR1 (top panels) or the HaDHFR^L22Y vectors (bottom panels). On the left, a sample of normal, freshly isolated C57BL/6 BM cells is shown with the SP gate indicated.