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B-hPD-1/hPD-L1/hCD40 mice
Strain Name

C57BL/6-Pdcd1tm1(PDCD1)Bcgen Cd274tm1(CD274)Bcgen 

CD40tm1(CD40)Bcgen/Bcgen

Common Name  B-hPD-1/hPD-L1/hCD40 mice
Background C57BL/6 Catalog number  131083
Related Genes 

PD-1(Programmed death-1) 

CD274 (CD274 antigen)

CD40 (CD40 antigen)
NCBI Gene ID
18566,60533,21939
Protein expression analysis

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Strain specific PD-1, PD-L1 and CD40 expression analysis in homozygous B-hPD-1/hPD-L1/hCD40 mice by flow cytometry. 

Splenocytes were collected from WT and homozygous B-hPD-1/hPD-L1/hCD40 (H/H) mice analyzed by flow cytometry with species-specific anti-PD-1 antibody. Mouse PD-1, PD-L1 and CD40 were detectable in WT mice. Human PD-1, PD-L1 and CD40 were exclusively detectable in homozygous B-hPD-1/hPD-L1/hCD40 but not WT mice. 


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Strain specific PD-1, PD-L1 and CD40 expression analysis in homozygous B-hPD-1/hPD-L1/hCD40 mice by flow cytometry. 

Splenocytes were collected from WT and homozygous B-hPD-1/hPD-L1/hCD40 (H/H) mice after stimulation with anti-CD3ε in vivo (7.5 μg/mice), and without stimulation, and analyzed by flow cytometry with species-specific anti-PD-1 antibody. Mouse PD-1, PD-L1 and CD40 were detectable in WT mice. Human PD-1, PD-L1 and CD40 were exclusively detectable in homozygous B-hPD-1/hPD-L1/hCD40 but not WT mice. After anti-CD3ε stimulation, hPD-1 protein expression was significantly increased.


Analysis of spleen leukocytes subpopulation in B-hPD-1/hPD-L1/hCD40 mice


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Analysis of spleen leukocyte subpopulations by FACS. 
Splenocytes were isolated from female C57BL/6 and B-hPD-1/hPD-L1/hCD40 mice (n=3, 7-week-old). Flow cytometry analysis of the splenocytes was performed to assess leukocyte subpopulations. A. Representative FACS plots. Single live cells were gated for the CD45+ population and used for further analysis as indicated here. B. Results of FACS analysis. Percent of T cells, B cells, NK cells, dendritic cells, granulocytes, monocytes and macrophages in homozygous B-hPD-1/hPD-L1/hCD40 mice were similar to those in the C57BL/6 mice, demonstrating that introduction of hPD-1/hPD-L1/hCD40 in place of their mouse counterpart does not change the overall development, differentiation or distribution of these cell types in spleen. Values are expressed as mean ± SEM.


Analysis of spleen T cell subpopulations in B-hPD-1/hPD-L1/hCD40 mice

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Analysis of spleen T cell subpopulations by FACS. 
Splenocytes were isolated from female C57BL/6 and B-hPD-1/hPD-L1/hCD40 mice(n=3, 7-week-old). Flow cytometry analysis of the splenocytes was performed to assess leukocyte subpopulations. A. Representative FACS plots. Single live CD45+ cells were gated for TCRβ+ T cell population and used for further analysis as indicated here. B. Results of FACS analysis. The percent of CD8+ T cells, CD4+ T cells, and Tregs in homozygous B-hPD-1/hPD-L1/hCD40 mice were similar to those in the C57BL/6 mice, demonstrating that introduction of hPD-1/hPD-L1/hCD40 in place of their mouse counterpart does not change the overall development, differentiation or distribution of these T cell subtypes in spleen. Values are expressed as mean ± SEM.


Analysis of lymph node leukocytes subpopulation in B-hPD-1/hPD-L1/hCD40 mice


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Analysis of lymph node leukocyte subpopulations by FACS. 
Lymph nodes were isolated from female C57BL/6 and B-hPD-1/hPD-L1/hCD40 mice(n=3, 7-week-old). Flow cytometry analysis of the splenocytes was performed to assess leukocyte subpopulations. A. Representative FACS plots. Single live cells were gated for the CD45+ population and used for further analysis as indicated here. B. Results of FACS analysis. Percent of T cells, B cells, NK cells, dendritic cells, granulocytes, monocytes and macrophages in homozygous B-hPD-1/hPD-L1/hCD40 mice were similar to those in the C57BL/6 mice, demonstrating that introduction of hPD-1/hPD-L1/hCD40 in place of their mouse counterpart does not change the overall development, differentiation or distribution of these cell types in lymph node. Values are expressed as mean ± SEM.


Analysis of lymph node T cell subpopulations in in B-hPD-1/hPD-L1/hCD40 mice



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Analysis of lymph node T cell subpopulations by FACS. 
Leukocytes were isolated from female C57BL/6 and B-hPD-1/hPD-L1/hCD40 mice(n=3, 7-week-old).  Flow cytometry analysis of the leukocytes was performed to assess leukocyte subpopulations. A. Representative FACS plots. Single live CD45+ cells were gated for TCRβ+ T cell population and used for further analysis as indicated here. B. Results of FACS analysis. The percent of CD8+ T cells, CD4+ T cells, and Tregs in homozygous B-hPD-1/hPD-L1/hCD40 mice were similar to those in the C57BL/6 mice, demonstrating that introduction of hPD-1/hPD-L1/hCD40 in place of their mouse counterpart does not change the overall development, differentiation or distribution of these T cell subtypes in lymph node. Values are expressed as mean ± SEM.

Analysis of blood leukocytes subpopulation in B-hPD-1/hPD-L1/hCD40 mice



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Analysis of blood leukocyte subpopulations by FACS
Blood cells were isolated from female C57BL/6 and B-hPD-1/hPD-L1/hCD40 mice(n=3, 7-week-old). Flow cytometry analysis of the splenocytes was performed to assess leukocyte subpopulations. A. Representative FACS plots. Single live cells were gated for the CD45+ population and used for further analysis as indicated here. B. Results of FACS analysis. Percent of T cells, B cells, NK cells, dendritic cells, granulocytes, monocytes and macrophages in homozygous B-hPD-1/hPD-L1/hCD40 mice were similar to those in the C57BL/6 mice, demonstrating that introduction of hPD-1/hPD-L1/hCD40 in place of their mouse counterpart does not change the overall development, differentiation or distribution of these cell types in blood. Values are expressed as mean ± SEM.

Analysis of blood T cell subpopulations in B-hPD-1/hPD-L1/hCD40 mice



from clipboard



Analysis of blood T cell subpopulations by FACS.
 Blood cells were isolated from female C57BL/6 and B-hPD-1/hPD-L1/hCD40 mice(n=3, 7-week-old).  Flow cytometry analysis of the splenocytes was performed to assess leukocyte subpopulations. A. Representative FACS plots. Single live CD45+ cells were gated for TCRβ+ T cell population and used for further analysis as indicated here. B. Results of FACS analysis. The percent of CD8+ T cells, CD4+ T cells, and Tregs in homozygous B-hPD-1/hPD-L1/hCD40 mice were similar to those in the C57BL/6 mice, demonstrating that introduction of hPD-1/hPD-L1/hCD40 in place of their mouse counterpart does not change the overall development, differentiation or distribution of these T cell subtypes in blood. Values are expressed as mean ± SEM.

Combination therapy of Pembrolizumab and Selicrelumab 

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Antitumor activity of of Pembrolizumab and Selicrelumab  in B-hPD-1/hPD-L1/hCD40 mice. (A) anti-human PD-1 antibody combined with anti-human CD40 antibody inhibited MC38-hPD-L1 tumor growth in B-hPD-1/hPD-L1/hCD40 mice. Murine colon cancer MC38-hPD-L1 cells (5×105) were subcutaneously implanted into homozygous B-hPD-1/hPD-L1/hCD40 mice(female, 7 week-old, n=5). Mice were grouped when tumor volume reached approximately 100-150 mm3, at which time they were treated with Pembrolizumab and Selicrelumab with doses and schedules indicated in panel A. (B) Body weight changes durin g treatment. As shown in panel A, combination of Pembrolizumab and Selicrelumab shows more inhibitory effects than individual groups, demonstrating that the B-hPD-1/hPD-L1/hCD40 mice provide a powerful preclinical model for in vivo evaluating combination therapy efficacy of PD-1 antibodies combined with anti-human CD40 antibodies. Values are expressed as mean ± SEM.(All antibodies were made in house)