Figure 7.

High-affinity TCR-mFc proteins bind peptide-pulsed antigen-presenting cells. a, wildtype (WT) RA14 and improved variants were expressed as TCR2ds–mFc proteins with mouse IgG2a hinge and Fc domains to reduce binding to human Fc receptors expressed on T2 cells. SDS-PAGE was used to assess protein size and purity (3 μg/lane). b, human T2 antigen-presenting cells were pulsed overnight with 100 μm NLV or KLV peptide and stained with 1 μm TCR2ds–mFc followed by 1:500 anti-mouse Fc-AF647 before flow cytometric analysis. This experiment was performed twice with similar results; representative data from one experiment are shown. c, human T2 antigen-presenting cells were pulsed overnight with NLV at 0, 0.5, 1, 5, 10, 25, 50, 100, 250, and 500 μm KLV peptide at 500 μm and stained with 1 μm α2.β8 in the TCR2dsΔgly-mFc format and 1:500 anti-mouse Fc-AF647 before flow cytometric analysis; 20,000 events were collected per condition to calculate a geometric mean fluorescence intensity (GMFI). Data shown are the average and range of two independent experiments performed with separate cells and protein preparations on different days.