Figure 2
Figure 2. Binding of E1, L1, and L2 to endogenous surface antigens on EBV-transformed BLCLs. (A) Representative histogram staining of HLA-A*0201+ and HLA-A*0201− donor samples with 3 of the TCR-like mAbs: E1 (blue), L1 (red), and L2 (green) before and after EBV infection and establishment of EBV BLCL outgrowth. (B) Normalized mean fluorescence intensity of the CD19+ B, CD3+ T, and CD56+ NK immune cell subsets in PBMCs with the 3 TCR-like mAbs. MFI values were normalized by dividing each MFI reading with their respective isotype controls. (C) nMFI of EBV BLCLs pre- and post-EBV immortalization. Blue and red symbols indicate HLA-A*0201+ and HLA-A*0201− samples, respectively. (D) Immunophenotyping summary of HLA-A*0201+ BLCLs examined in this study. Normalized MFI value of 0 to less than 2 indicates negative staining (−), above 2 to 9.99 (+), 10 to 99.9 (++), and 100 to 1000 (+++). (E) Compilation of normalized mean fluorescence intensity values of each HLA-A*0201+ BLCL from this study, with HLA-A2+ EBV− BJAB (open square) and HLA-A2− EBV+ Raji (open circle) cell lines as controls. Unpaired Student t test was performed to compare the binding profiles between each TCR-like mAb on HLA-A*0201+ BLCLs. **P < .001; ***P < .0001. (F) Confocal microscope images of BLCLs stained with the respective TCR-like mAbs (R-phycoerythrin), β-2-microglobulin (AlexaFluor-488), and Hoechst dye (blue). Scale bars, 10 μm. (G) Immunoblot detection of EBV latent proteins in 2 HLA-A*0201+ BLCLs AM003, CM462, HLA-A*0201− atypical latency III cell line Raji, and EBV− BJAB as a control. GAPDH served as a loading control. (H) Representative histograms staining indicating BJAB as being HLA-A2+ and Raji cell line as HLA-A2−, with TCR-like mAbs staining profiles similar to that of isotype and unstained controls.

Binding of E1, L1, and L2 to endogenous surface antigens on EBV-transformed BLCLs. (A) Representative histogram staining of HLA-A*0201+ and HLA-A*0201 donor samples with 3 of the TCR-like mAbs: E1 (blue), L1 (red), and L2 (green) before and after EBV infection and establishment of EBV BLCL outgrowth. (B) Normalized mean fluorescence intensity of the CD19+ B, CD3+ T, and CD56+ NK immune cell subsets in PBMCs with the 3 TCR-like mAbs. MFI values were normalized by dividing each MFI reading with their respective isotype controls. (C) nMFI of EBV BLCLs pre- and post-EBV immortalization. Blue and red symbols indicate HLA-A*0201+ and HLA-A*0201 samples, respectively. (D) Immunophenotyping summary of HLA-A*0201+ BLCLs examined in this study. Normalized MFI value of 0 to less than 2 indicates negative staining (−), above 2 to 9.99 (+), 10 to 99.9 (++), and 100 to 1000 (+++). (E) Compilation of normalized mean fluorescence intensity values of each HLA-A*0201+ BLCL from this study, with HLA-A2+ EBV BJAB (open square) and HLA-A2 EBV+ Raji (open circle) cell lines as controls. Unpaired Student t test was performed to compare the binding profiles between each TCR-like mAb on HLA-A*0201+ BLCLs. **P < .001; ***P < .0001. (F) Confocal microscope images of BLCLs stained with the respective TCR-like mAbs (R-phycoerythrin), β-2-microglobulin (AlexaFluor-488), and Hoechst dye (blue). Scale bars, 10 μm. (G) Immunoblot detection of EBV latent proteins in 2 HLA-A*0201+ BLCLs AM003, CM462, HLA-A*0201 atypical latency III cell line Raji, and EBV BJAB as a control. GAPDH served as a loading control. (H) Representative histograms staining indicating BJAB as being HLA-A2+ and Raji cell line as HLA-A2, with TCR-like mAbs staining profiles similar to that of isotype and unstained controls.

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