Basic Information
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Targeting strategy
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Gene targeting strategy for B-hGAS6 mice. The exons 1-15 of mouse Gas6 gene that encode the full-length protein were replaced by human GAS6 exons 1-15 in B-hGAS6 mice.
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mRNA expression analysis
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Strain specific analysis of GAS6 gene expression in wild-type C57BL/6 mice and B-hGAS6 mice by RT-PCR. Mouse Gas6 mRNA was detectable only in ovary of wild-type C57BL/6 mice (+/+). Human GAS6 mRNA was detectable only in homozygous B-hGAS6 mice (H/H), but not in wild-type mice.
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Protein expression analysis
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Strain specific GAS6 expression analysis in homozygous B-hGAS6 mice by ELISA. Plasma was collected from wild-type mice (+/+) and homozygous B-hGAS6 mice (H/H), and analyzed by ELISA with species-specific GAS6 ELISA kit. Mouse GAS6 was detectable only in wild-type mice. Human GAS6 was exclusively detectable in homozygous B-hGAS6 mice but not in wild-type mice.
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Analysis of leukocytes cell subpopulation in spleen
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Analysis of spleen leukocyte subpopulations by FACS. Splenocytes were isolated from female C57BL/6 and B-hGAS6 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/B cells, NK cells, dendritic cells, granulocytes, monocytes and macrophages in homozygous B-hGAS6 mice were similar to those in the C57BL/6 mice, demonstrating that GAS6 humanized does not change the overall development, differentiation or distribution of these cell types in spleen. Values are expressed as mean ± SEM.
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Analysis of T cell subpopulation in spleen
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Analysis of spleen T cell subpopulations by FACS. Splenocytes were isolated from female C57BL/6 and B-hGAS6 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 CD8+ T cells and CD4+ T cells in homozygous B-GAS6 mice were similar to those in the C57BL/6 mice, demonstrating that introduction of hGAS6 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.
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Analysis of leukocytes cell subpopulation in lymph node
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Analysis of lymph node leukocyte subpopulations by FACS. Leukocytes were isolated from female C57BL/6 and B-hGAS6 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 CD45+ population and used for further analysis as indicated here. B. Results of FACS analysis Percent of T cells, B cells, NK cells, in homozygous B-hGAS6 mice were similar to those in the C57BL/6 mice, demonstrating that introduction of hGAS6 in place of its mouse counterpart does not change the overall development, differentiation or distribution of these cell types in lymph node. Values are expressed as mean ± SEM.
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Analysis of T cell subpopulation lymph node
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Analysis of lymph node leukocyte subpopulations by FACS. Leukocytes were isolated from female C57BL/6 and B-hGAS6 mice (n=3, 6-week-old). Flow cytometry analysis of the leukocytes was performed to assess leukocyte subpopulations. 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 Treg cells in homozygous B-hGAS6 mice were similar to those in the C57BL/6 mice, demonstrating that introduction of hGAS6 in place of its mouse counterpart does not change the overall development, differentiation or distribution of these cell types in lymph node. Values are expressed as mean ± SEM.
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Analysis of leukocytes cell subpopulation in blood
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Analysis of blood leukocyte subpopulations by FACS. Blood were isolated from female C57BL/6 and B-hGAS6 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 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-hGAS6 mice were similar to those in the C57BL/6 mice, demonstrating that introduction of hGAS6 in place of its mouse counterpart does not change the overall development, differentiation or distribution of these cell types in blood. Values are expressed as mean ± SEM.
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Analysis of T cell subpopulation in blood
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Analysis of blood leukocyte subpopulations by FACS. Blood were isolated from female C57BL/6 and B-hGAS6 mice (n=3, 6-week-old). Flow cytometry analysis of the leukocytes was performed to assess leukocyte subpopulations. 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+, CD8+, Treg cells in homozygous B-hGAS6 mice were similar to those in the C57BL/6 mice, demonstrating that introduction of hGAS6 in place of its mouse counterpart does not change the overall development, differentiation or distribution of these cell types in blood. Values are expressed as mean ± SEM.
