• CD3 consists of four protein chains (CD3E, CD3D, CD3G and CD3Z), which are important biological markers on the T cell membrane. CD3 can form a TCR/CD3 complex with the T cell receptor, participating in the regulation of T cell antigen recognition, signal transduction and T cell development. CD3 antibodies can mimic antigen signals and ​​non-specifically​​ activate all T cells in the tumor microenvironment. Activated T cells highly express PD-1. When PD-1 binds to PD-L1 on the surface of tumor cells, it transmits an inhibitory signal into the T cell, causing it to become inactivated, exhausted, and unable to kill tumor cells. The role of PD-1 antibodies is to ​​block this "brake" signal​​.
• B-hPD-1 plus/hPD-L1/hCD3EDG mice were obtained by mating B-hPD-1 plus/hPD-L1 mice (112550) and B-hCD3EDG mice (110039). Human PD-1 and PD-L1 were exclusively detectable on T cell subgroups of homozygous B-hPD-1 plus/hPD-L1/hCD3EDG mice but not in C57BL/6 mice. Human CD3E was exclusively detectable on T cells of homozygous B-hPD-1 plus/hPD-L1/hCD3EDG mice but not in C57BL/6 mice. Humanization of PD-1, PD-L1 and CD3EDG does not change the overall frequency or distribution of immune cell types in spleen, blood, and lymph nodes.
• This product is used for the combination therapy of PD-1/PD-L1 inhibitors and CD3-targeted drugs.

Strain specific PD-1 expression analysis in homozygous B-hPD-1 plus/hPD-L1/hCD3EDG mice by flow cytometry. Splenocytes were collected from C57BL/6 mice and homozygous B-hPD-1 plus/hPD-L1/hCD3EDG mice, either untreated (upper panel) or treated (lower panel) with anti-CD3ε in vivo (7.5 μg per mouse, 24 hours, i.p.). Cells were analyzed by flow cytometry using species-specific antibodies: anti-mouse PD-1 (Biolegend, 109104) and anti-human PD-1 (Biolegend, 329908). Mouse PD-1 was detectable in C57BL/6 mice, while human PD-1 was exclusively detectable in homozygous B-hPD-1 plus/hPD-L1/hCD3EDG mice but not in C57BL/6 mice.

Strain specific PD-L1 expression analysis in homozygous B-hPD-1 plus/hPD-L1/hCD3EDG mice by flow cytometry. Splenocytes were collected from C57BL/6 mice and homozygous B-hPD-1 plus/hPD-L1/hCD3EDG mice, either untreated (upper panel) or treated (lower panel) with anti-CD3ε in vivo (7.5 μg per mouse, 24 hours, i.p.). Cells were analyzed by flow cytometry using species-specific antibodies: anti-mouse PD-L1 (Biolegend, 124312) and anti-human PD-L1 (Biolegend, 329706). Mouse PD-L1 was detectable in C57BL/6 mice, while human PD-L1 was exclusively detectable in homozygous B-hPD-1 plus/hPD-L1/hCD3EDG mice but not in C57BL/6 mice.

Strain specific CD3E expression analysis in homozygous B-hPD-1 plus/hPD-L1/hCD3EDG mice by flow cytometry. Splenocytes were collected from C57BL/6 mice and homozygous B-hPD-1 plus/hPD-L1/hCD3EDG mice, and analyzed by flow cytometry with species-specific anti-mouse CD3E antibody (Biolegend, 100321), and anti-human CD3E antibody (BD, 562426). Mouse CD3E was only detectable in wild-type C57BL/6 mice. Human CD3E was exclusively detectable in homozygous B-hPD-1 plus/hPD-L1/hCD3EDG mice.

Frequency of leukocyte subpopulations in spleen by flow cytometry. Splenocytes were isolated from wild-type C57BL/6 mice (female, n=3, 8-week-old) and homozygous B-hPD-1 plus/hPD-L1/hCD3EDG mice (female, n=3, 8-week-old). A. Flow cytometry analysis of the splenocytes was performed to assess the frequency of leukocyte subpopulations. B. Frequency of T cell subpopulations. Percentages of T cells, B cells, NK cells, dendritic cells, neutrophils, monocytes, macrophages, CD4+ T cells, CD8+ T cells and Tregs in B-hPD-1 plus/hPD-L1/hCD3EDG mice were similar to those in C57BL/6 mice. The same results were observed in lymph nodes and blood. Data were not shown. Values were expressed as mean ± SEM. Significance was determined by two-way ANOVA test.  *P < 0.05, **P < 0.01, ***p < 0.001.