Cells transfected with scTCR recognize MUC1-positive tumors and synthetic MUC1 antigen. (A) Mammalian expression vector encoding MA scTCR gene. The scTCR was cloned into the pEF6 vector. BGH indicates bovine growth hormone; EF-α, elongation factor α. (B) Cell surface expression of the scTCR in RBL cells or (C) BWZ cells stably transfected with the scTCR-pEF6 vector. Cells were stained with anti-TCR βF1 (open histogram) or with isotype control (filled histogram) antibody. (D) Degranulation of RBL cells or RBL-scTCR cells following stimulation with platebound βF1 antibody or MUC1 140-mer peptide. Specific degranulation is presented as percent of maximum degranulation induced by TCR cross-linking with βF1 antibody. (E) IL-2 enzyme-linked immunosorbent assay (ELISA) for BWZ or BWZ-scTCR following stimulation with platebound anti-TCR βF1 antibody, ionomycin plus PMA (I/P), DM6 (MUC1-negative tumor), HPAF, or T3M4 (MUC1-positive tumors). IL-2 in culture supernatant was measured by ELISA, and values were plotted on the y-axis as picograms per milliliter. Cells were stimulated as indicated.