Fig. 1. SIG35α but not DMF5α expressing SupT1 cells are stained by A2/MART1 multimers when paired with the endogenous irrelevant TCRβ chain of SupT1 cells.

The human TCRα⁻/β⁺ pre-T cell leukemia cell line, SupT1 was transduced with 5 different clonotypic A2/MART1-specific TCRα chains, SIG35α/ΔNGFR, DMF5α/ΔNGFR, SIG35αN/ΔNGFR, or DMF5αS/ΔNGFR, or TCRαβ chains, DMF5αβ/ΔNGFR. SIG35αN is a SIG35α-derived mutant encoding Asn instead of Ser at the V-J junction. DMF5 is a high affinity A2/MART1 TCR (48). DMF5αS is a DMF5α-derived mutant coding for Ser instead of Asn at the V-J junction. All TCRα genes were fused with ΔNGFR gene via an optimized intervening sequence consisting of a furin cleavage site, an SGSG spacer sequence, and an F2A sequence (35). ΔNGFR alone was employed as a control. The transduction efficiency of SupT1 transfectants was approximately 90% as determined by the percentage of ΔNGFR⁺ cells (data not shown). All SupT1 transfectants were stained with A2/MART1 or A2/HIV multimer along with anti-CD3 mAb. Data shown are gated on ΔNGFR⁺ cells and a representative of two independent experiments.