Fig. 2

Analysis of Assay Dynamic Range using Transduced T cells and Lentiviral Vector. T cells engineered to express an HPV-16 E7 oncogene-specific TCR and the lentiviral vector used to produce these cells were used to assess the dynamic range of the ddPCR vector copy number assay. a HPV16 E7 TCR engineered T cells were serial diluted twofold with untransduced lymphocytes and percentage of TCR-beta positive cells was determined by flow cytometry. HPV16 E7 TCR engineered cells that were not diluted are shown in panel a, cells diluted 1:2 are shown in panel b, 1:4 diluted in panel c, 1:8 diluted cells in panel d, and 1:16 diluted in panel e. Flow cytometry analysis of untransduced lymphocytes are show in panel f. The representative flow plots were gated on viable CD3⁺ T cells. b For each of the HPV16 E7 TCR engineered T cell samples serially diluted with untransduced lymphocytes, the log of the transduction efficiency and the log of the dilution factor is shown. The line of best fit with confidence intervals are shown. Each point represents the average of two measurements. c The HPV16 E7 TCR engineered T cell samples were also analyzed for vector copy number by ddPCR and the log of the vector copy number and dilution for each sample is shown. Each point represents the average of two measurements. The line of best fit with confidence intervals is shown. d The HPV-16 E7 TCR lentiviral vector was diluted with DNA from untransduced lymphocytes cells and evaluated by ddPCR and the log of the vector copy number and dilution for each sample is shown. Each point represents the average of two measurement. The line of best fit with confidence intervals is shown