B-hPD-L1 low B16-F10

Basic Information

Targeting strategy

Gene targeting strategy for B-hPD-L1 low B16-F10 cells. The exogenous promoter and human PD-L1 coding sequence was inserted to replace part of murine exon 3. The insertion disrupts the endogenous murine Pdl1 gene, resulting in a non-functional transcript.

Protein expression analysis

PD-L1 expression analysis in B-hPD-L1 low B16-F10 cells by flow cytometry. Single cell suspensions from wild-type B16-F10, B-hPD-L1 B16-F10 and B-hPD-L1 low B16-F10 cultures were stained with species-specific anti-PD-L1 antibody. Mouse PD-L1 was detectable in wild-type B16-F10 cells. Human PD-L1 was detectable in B-hPD-L1 low B16-F10 cells, and human PD-L1 was expressed highly on the surface of B-hPD-L1 B16-F10 cells. The 2-G02 clone of B-hPD-L1 low B16-F10 cells was used for in vivo experiments.

Tumor growth curve & Body weight changes

Subcutaneous homograft tumor growth of B-hPD-L1 low B16-F10 cells. Wild-type B16-F10, B-hPD-L1 B16-F10 and B-hPD-L1 low B16-F10 cells (2×10⁵) were subcutaneously implanted into C57BL/6 mice (female, 7-8-week-old, n=5). Tumor volume and body weight were measured twice a week. (A) Average tumor volume ± SEM. (B)  Body weight (Mean± SEM). Volume was expressed in mm³ using the formula: V=0.5 X long diameter X short diameter². As shown in panel A, B-hPD-L1 low B16-F10 cells were able to establish tumors in vivo and can be used for efficacy studies.

Protein expression analysis of tumor cells

B-hPD-L1 low B16-F10 and B-hPD-L1 B16-F10 cells were subcutaneously transplanted into C57BL/6 mice (n=5), and on 20 days post inoculation, tumor cells were harvested and assessed for human PD-L1 expression by flow cytometry. As shown, human PD-L1 was expressed on the surface of B-hPD-L1 low B16-F10 tumor cells, and it was expressed highly on the surface of B-hPD-L1 B16-F10 tumor cells. Therefore, B-hPD-L1 low B16-F10 cells can be used for in vivo efficacy studies of novel PD-L1 therapeutics.

FMO Control: tumor cells only stained anti-hPD-L1 antibody.