Basic Information

Strain Name
C57BL/6-Tnfsf11tm1(TNFSF11)/Bcgen
Stock Number
111072
Common Name
B-hRANKL mice
Source/Investigator
Bcgen (Beijing Biocytogen Co., Ltd)
Aliases
CD254, ODF, OPGL, OPTB2, RANKL, TNLG6B, TRANCE, hRANKL2, sOdf
Species
C57BL/6
Appearance
Black
Genotypes
Homozygous
NCBI Gene ID

Targeting strategy

Gene targeting strategy for B-hRANKL mice. The exons 4~5 of mouse Rankl gene that encode the extracellular region were replaced with human RANKL exons 4~5 in B-hRANKL mice.

Details

mRNA expression analysis

Strain specific analysis of RANKL gene expression in wild-type C57BL/6 mice and B-hRANKL mice by RT-PCR. Mouse Rankl mRNA was detectable only in thymocytes of wild-type C57BL/6 mice (+/+). Human RANKL mRNA was detectable only in homozygous B-hRANKL mice but not in wild-type mice.

Protein expression analysis

Strain specific RANKL expression analysis in wild-type C57BL/6 mice and homozygous B-hRANKL mice by flow cytometry. CD4+ T cells were isolated from the spleen of wild-type C57BL/6 mice (+/+) and homozygous B-hRANKL mice (H/H) stimulated with anti-mCD3e and anti-mCD28 antibodies in vitro. Mouse RANKL was detectable only in wild-type C57BL/6 mice. Human RANKL was exclusively detectable in homozygous B-hRANKL mice.

Analysis of leukocytes cell subpopulation in spleen

Analysis of spleen leukocyte subpopulations by FACS. Splenocytes were isolated from female C57BL/6 and B-hRANKL mice (n=3, 6-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-hRANKL mice were similar to those in the C57BL/6 mice, demonstrating that introduction of hRANKL in place of its 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 T cell subpopulation in spleen

Analysis of spleen T cell subpopulations by FACS. Splenocytes were isolated from female C57BL/6 and B-hRANKL mice (n=3, 6-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 CD4+ T cells, CD8+ T cells and Tregs in homozygous B-hRANKL mice were similar to those in the C57BL/6 mice, demonstrating that introduction of hRANKL in place of its 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 leukocytes cell subpopulation in lymph node

Analysis of lymph node leukocyte subpopulations by FACS. Lymph nodes were isolated from female C57BL/6 and B-hRANKL mice (n=3, 6-week-old). Flow cytometry analysis of the leukocytes 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 and NK cells in homozygous B-hRANKL mice were similar to those in the C57BL/6 mice, demonstrating that introduction of hRANKL in place of its 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 T cell subpopulation in lymph node

Analysis of lymph node T cell subpopulations by FACS. Leukocytes were isolated from female C57BL/6 and B-hRANKL mice (n=3, 6-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 CD4+ T cells, CD8+ T cells, and Tregs in homozygous B-hRANKL mice were similar to those in the C57BL/6 mice, demonstrating that introduction of hRANKL in place of its 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 leukocytes cell subpopulation in blood

Analysis of blood leukocyte subpopulations by FACS. Blood cells were isolated from female C57BL/6 and B-hRANKL mice (n=3, 6-week-old). Flow cytometry analysis of the blood cells 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-hRANKL mice were similar to those in the C57BL/6 mice, demonstrating that introduction of hRANKL in place of its mouse counterpart does not change the overall development, differentiation or distribution of these T cell subtypes in blood. Values are expressed as mean ± SEM.

Analysis of T cell subpopulation in blood

Analysis of blood T cell subpopulations by FACS. Blood cells were isolated from female C57BL/6 and B-hRANKL mice (n=3, 6-week-old). Flow cytometry analysis of the blood cells 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 CD4+ T cells, CD8+ T cells, and Tregs in homozygous B-hRANKL mice were similar to those in the C57BL/6 mice, demonstrating that introduction of hRANKL in place of its mouse counterpart does not change the overall development, differentiation or distribution of these T cell subtypes in blood. Values are expressed as mean ± SEM.

Summary

mRNA expression analysis:
Mouse Rankl mRNA was detectable only in thymocytes of wild-type C57BL/6 mice (+/+). Human RANKL mRNA was detectable only in homozygous B-hRANKL mice.

Protein expression analysis:
Mouse RANKL was detectable only in wild-type C57BL/6 mice. Human RANKL was exclusively detectable in homozygous B-hRANKL mice.

Leukocytes cell subpopulation analysis:
RANKL humanized does not change the overall development, differentiation or distribution of immune cell types in spleen, lymph nodes and blood.