Journal: Science Advances
Article Title: Cell surface–tethered IL-12 repolarizes the tumor immune microenvironment to enhance the efficacy of adoptive T cell therapy
doi: 10.1126/sciadv.abi8075
Figure Lengend Snippet: ( A and B ) Quantification of the number of receptor-targeted antibodies bound per cell (A) or percent of cells expressing the given receptor (B) on resting (gray) or CD3/CD28-stimulated (blue) T cells. In (A), only the expressing population of cells is included for analysis; for example, CD8-negative cells are excluded from analysis of CD8 antibody binding capacity in (A). Inset: Antibodies per cell for cytokine and checkpoint receptors with enlarged y axis. Error bars represent SD for T cells from three human donors. Multiple antibody clones targeting the same receptor were used when available to improve accuracy. ( C ) Fold difference of the average number of antibodies bound per cell for the abundant receptors versus cytokine and checkpoint receptors. Antibody clones are in the same order as in (A) and (B). ( D to I ) Construct design (D and E), dose-dependent loading (F and G), and relative cytokine loading at 500 nM condition (H and I) for surface-tethered IL-12 and IL-15 constructs. Variable domains for anti-CD11a, anti-CD18, and anti-CD45 antibodies were from antibody clones MHM24, 1B4, and 9.4, respectively. PD-1, programmed cell death protein 1; MHC, major histocompatibility complex; MFI, median fluorescent intensity; Ab, antibody.
Article Snippet: A standard curve was generated using fluorescent bead standards (Quantum MESF Alexa 647 Beads, Bangs Laboratories Inc.) to correlate fluorochrome median fluorescent intensity (MFI) to the number of fluorochromes.
Techniques: Expressing, Binding Assay, Clone Assay, Construct, Immunopeptidomics