Supplemental Figure 1 A B C Mock MOI-10 MOI % 21.8% 79.7%

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Supplemental Figure 1 A B C Mock MOI-10 MOI-100 0.5% 21.8% 79.7% CD3 PerCP NY-ESO-1 Tetramer PE B Supplemental Figure 1. Validation of sr39TK function in Jurkat cells. (A) Mock or ESO/TK transduced Jurkats were evaluated by flow cytometry for NY-ESO-1 tetramer binding. (B) Cells were cultured with 0.5µCi [18F]-FHBG for 1 hour, washed, and evaluated for uptake. (C) Cells were cultured in half-log increasing concentrations of GCV for 48h and evaluated for viability. C

Supplemental Figure 2 Thymus L Humerus Muscle L Femur R Humerus R Femur A Supplemental Figure 2. ROI Analysis. (A) Schematic of ROIs drawn on PET imaged mice. 3d cylindrical or spherical ROIs were drawn using Amide software on hematopoietic niches to determine probe uptake and arm muscle for background subtraction. (B) There was significant [18F]-FHBG uptake in the hematopoietic niches of ESO/TK mice compared with mock transduced controls. (C) [18F]-FHBG uptake in non-transduced humanized mice before and after GCV treatment. B C P < 0.0001

Supplemental Figure 3 A B Supplemental Figure 3. Chimerism, vector marking, and PET probe signal in vertebrae. (A) Cells harvested from individual vertebrae were evaluated for human chimerism by flow cytometry and vector marking by qPCR. (B) Data were matched with [18F]-FHBG PET scan.