Figure 1. Generation and validation of PDL1-K562 cells. (A) Schematic representation of the lentiviral vector used to overexpress the human CD274 gene (GenBank accession number: NM_014143). (B) Detection of mCherry fluorescent protein expression in PDL1-K562 cells by fluorescence microscopy (100×). (C) Western blotting detection of PD-L1 in K562 cells. A 48-55 kDa protein band consistent with glycosylated PD-L1 was identified. Lanes 1-3, anti-flag antibody; Lanes 4-5, anti-PDL1 antibody; Lane 1, SURVIVIN-3FLAG-EGFP (48 kDa); Lanes 2 and 4, K562 cell lysates; Lanes 3 and 5, PDL1-K562 cell lysates. (D) Flow cytometry data showing that PD-L1 is expressed on the surface of almost all PDL1-K562 cells.