Gene targeting strategy for B-CAG-hPD-L1 Raji-luc. The human full-length CD274 CDS region with CAG promoter was inserted in the AAVS1 locus exon1 of B-luc-GFP Raji.
Protein expression analysis
The PD-L1 expression analysis in B-CAG-hPD-L1 Raji-luc cells by flow cytometry. Single cells were collected from B-luciferase-GFP Raji-Luc and B-CAG-hPD-L1 Raji-luc cells, and analyzed by flow cytometry with species-specific anti-PD-L1 antibody. Human PD-L1 was exclusively detectable in B-CAG-hPD-L1 Raji-luc cells but not B-luc-GFP Raji cells.
Tumor Growth Curve & Body Weight Changes
Subcutaneous xenograft tumor growth of B-CAG-hPD-L1 Raji-luc cells. B-CAG-hPD-L1 Raji-luc cells and B-luc-GFP Raji cells (5×105) were subcutaneously implanted into the B-NDG mice (n=?).Tumor size and mice body weight were measured twice a week. (A) Tumor average volume ± SEM, (B) Mice body weight (Mean± SEM). Volume was expressed in mm3 using the formula: V=0.5a X b2, where a and b were the long and short diameters of the tumor, respectively. As shown in panel A, B-CAG-hPD-L1 Raji-luc were able to establish tumor in vivo and can be used for efficacy study.
Protein expression analysis in tumor cells
B-CAG-hPD-L1 Raji-luc cells were transplanted into B-NDG mice, and on ? days post inoculation, certain volumes of tumors as was taken to detect the expression level of the cells. As Shown,human PD-L1 was highly expressed on the surface of tumor cells. B-CAG-hPD-L1 Raji-luc can be used for in vivo efficacy validation.
In vivo Imaging Analysis
Tumor growth and in vivo imaging of B-CAG-hPD-L1 Raji-luc cells. B-CAG-hPD-L1 Raji-luc and B-luciferase-GFP Raji-Luc cells(5×105) were injected by tail vein into B-NDG mice (n=？). Signal intensity and body weight were measured twice a week. (A) Imaging was performed on days 0, 4, 6, 8, 11, 15, and 18，(B) Mice body weight (Mean ± SEM). B-CAG-hPD-L1 Raji-luc cells can be used for in vivo efficacy evaluation.