Figure 4.
Figure 4. In vivo selection of GFP+ peripheral blood cells in monkey 96E113 following drug treatment. (A) Vector DNA copy number by PCR 30 days before drug treatment. DNA from Ficoll-purified granulocytes (Gran) and mononuclear cells (Mon) that was obtained from peripheral blood samples 30 days prior to drug treatment was analyzed for sequences from the MGirL22Y vector (top row) or endogenous β-actin genes (bottom row). As a negative control, whole PB DNA from an animal that did not receive a transplant (Untx) was assayed. Copy number controls represent serial dilutions of a single copy vector producer cell line, with the indicated percentage of vector copy per cell shown above each column. The calculated percentage of copy numbers, normalized for actin signal, are shown below each experimental sample. (B) Myelosuppression following an 8-day course with peg-SCF and G-CSF given concurrently with TMTX 6 mg/kg/d × 5 and NBMPR-P 3 mg/kg/d × 5. As before, baseline and posttreatment values are shown for the absolute neutrophil count and the platelet count. The axes are labeled as in Figure 3A. (C) Changes in the percentage of GFP-positive peripheral blood cells prior to, during, and following drug treatment. As indicated by the legend, flow cytometry measurements for GFP are shown for granulocytes, monocytes, and lymphocytes. Note the maximum value of the y-axis is 70%. All days are relative to the first day of drug treatment. (D) Representative flow cytometry data for granulocytes, monocytes, and lymphocytes obtained at baseline and at various days relative to the first day of treatment. Percent GFP+ cells are shown for each histogram.

In vivo selection of GFP+ peripheral blood cells in monkey 96E113 following drug treatment. (A) Vector DNA copy number by PCR 30 days before drug treatment. DNA from Ficoll-purified granulocytes (Gran) and mononuclear cells (Mon) that was obtained from peripheral blood samples 30 days prior to drug treatment was analyzed for sequences from the MGirL22Y vector (top row) or endogenous β-actin genes (bottom row). As a negative control, whole PB DNA from an animal that did not receive a transplant (Untx) was assayed. Copy number controls represent serial dilutions of a single copy vector producer cell line, with the indicated percentage of vector copy per cell shown above each column. The calculated percentage of copy numbers, normalized for actin signal, are shown below each experimental sample. (B) Myelosuppression following an 8-day course with peg-SCF and G-CSF given concurrently with TMTX 6 mg/kg/d × 5 and NBMPR-P 3 mg/kg/d × 5. As before, baseline and posttreatment values are shown for the absolute neutrophil count and the platelet count. The axes are labeled as in Figure 3A. (C) Changes in the percentage of GFP-positive peripheral blood cells prior to, during, and following drug treatment. As indicated by the legend, flow cytometry measurements for GFP are shown for granulocytes, monocytes, and lymphocytes. Note the maximum value of the y-axis is 70%. All days are relative to the first day of drug treatment. (D) Representative flow cytometry data for granulocytes, monocytes, and lymphocytes obtained at baseline and at various days relative to the first day of treatment. Percent GFP+ cells are shown for each histogram.

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