Figure 6.

Generation of a cocal producer cell line for WT1-specific TCR LV. (a) Lentiviral vector (LV) titer measured by quantitative PCR (qPCR) in single cocal T-cell receptor (TCR) producer clones (open circles) or bulk cells (bar). Unconcentrated titer is given as infectious units/ml (IU/ml). (b) Unconcentrated (1×) and concentrated (100×) LV titer measured by qPCR from multiple experiments (n = 5) using the best producer clone identified in a. Bar graph shows mean titer and error bars show SE of the mean. (c) Stability of cocal TCR LV producer cell line based on unconcentrated titer after long-term culture. (d) TCR expression measured in H9 cell line transduced with different amounts of LV made with the standard protocol or with the producer cell line. TCR expression was detected by surface antibody staining at 3 days post transduction and the fraction of TCR positive cells was quantified by flow cytometry. (e) TCR expression measured in human CD4⁺ cells transduced with different amounts of vector as described in d. (f) Histogram showing levels of TCR expression measured in live human CD4⁺ cells from e transduced with 5μl LV made with producer cells (gray line) or with standard conditions (black line), as compared to untransduced/stained cells (red line) and to unstained cells (gray shade).