(A) CD34 cells were transduced with vCWRHIVAPAP at MOI of 3 (particle MOI, 600) immediately after isolation and placed in culture. At the indicated times suspension cells were harvested and placed in media containing increased concentrations of IL-3, IL-6, and stem cell factor to boost the mitotic index before blocking with colcemid as described in Materials and Methods. Metaphases were identified and scored for the presence of vector-specific signal. Only metaphases showing evidence of vector signal on both sister chromatids were scored as positive. Untransduced controls never hybridized with the probe in every CD34 sample analyzed. PLAP expression from the RSV promoter in vCWRHIVAPAP was analyzed concurrently with FISH where indicated. (B) Flow-sorted CD34⁺38⁻ cells were transduced with vCWRHIVAPAP at functional MOIs of 1 to 3. Cells were harvested from cultures at the indicated time points and assayed by FISH using a vector-specific probe as described above. Cells donors KL and NT were transduced in parallel.

Abbreviation: —, not done.

LTC-ICs were also analyzed from these donors (Table 1).

In this sample, most CD34 cells had differentiated into adherent macrophages by 9 weeks and few suspension cells were available for analyses.