Cotransfections with fXI/PKA2. (A) Comparison of wild-type (WT) and fXI/PKA2 expression. The 293 cells were transfected with the following combinations of expression constructs: (1) 0.08 μg WT-fXI; (2) 0.08 μg fXI/PKA2; (3) 0.16 μg WT-fXI; (4) 0.08 μg fXI/PKA2 + 0.08 μg WT-fXI; and (5) 0.16 μg pJVCMV. Shown are relative concentrations of protein in media 67 hours after transfection compared with the mean for wild-type fXI (assigned a value of 100%). Results are means and standard deviations for 6 separate transfections, each tested in triplicate by ELISA and corrected for transfection efficiency by Renilla luciferase assay. Capture antibody was polyclonal (▪) or monoclonal 1A6 (□) anti-fXI antibody; 1A6 recognizes the fXI A2 domain and will not recognize fXI/PKA2. (B) Cotransfection of fXI/PKA2 with mutant fXI constructs. Transient transfections of 293 cells were performed with 0.08 μg fXI/PKA2 expression construct and 0.08 μg of (1) pJVCMV; (2) fXI-Gly400Val; and (3) fXI-Trp569Ser. Lane 4 represents control transfection with 0.16 μg pJVCMV. Shown are relative concentrations of protein in media 67 hours after transfection compared with the mean for fXI/PKA2, which was assigned a value of 100%. Results are means and standard deviations for 6 separate transfections, each tested in triplicate by ELISA and corrected for transfection efficiency by Renilla luciferase assay. Capture antibody was polyclonal (▪) or monoclonal 1A6 (□) anti-fXI antibody.