• 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. CD20 is a marker of B cells, and it begins to express in Pre-B cells and loses expression after differentiating into plasma cells. CD20 is expressed on the surface of normal and about 95% malignant B lymphocytes, but not expressed in hematopoietic stem cells, plasma cells, or other normal tissues. CD20 is regarded as an ideal target for the treatment of B-cell lymphoma and autoimmune diseases.
• B-hCD3E/hCD20 mice were obtained by mating B-hCD3E mice(110008) and B-hCD20 mice(111231). In this mice, the exons 2-6 of mouse Cd3e gene that encode the extracellular domain were replaced by human CD3E exons 2-7. The exons 2-7 of mouse Cd20 gene that encode the whole molecule (ATG to STOP codon) are replaced by human counterparts in B-hCD3E/hCD20 mice.
• Human CD3E can be detected on T cells from spleen of homozygous B-hCD3E/hCD20 mice, but not in wild-type mice. Human CD20 can be detected on B cells from spleen of homozygous B-hCD3E/hCD20 mice. Following humanization of CD3E and CD20, the percentages of T cells, B cells, NK cells, dendritic cells, neutrophils, monocytes and macrophages in B-hCD3E/hCD20 mice were similar to those wild-type mice.
• This product is used for the pharmacological and safety evaluation of therapeutic drugs, such as anti-CD20 antibodies, bi-specific antibodies, CAR-T cell therapies, and other drugs. Disease areas include oncology and autoimmune diseases.

Gene targeting strategy for B-hCD3E/hCD20 mice.
The exons 2-6 of mouse Cd3e gene that encode the extracellular domain were replaced by human CD3E exons 2-7 in B-hCD3E/hCD20 mice. The exons 2-7 of mouse Cd20 gene that encode the full-length protein were replaced by human CD20 exons 2-7 in B-hCD3E/hCD20 mice.

Strain specific CD3E expression analysis in homozygous B-hCD3E/hCD20 mice by flow cytometry.
Splenocytes were collected from wild type (WT) mice (+/+) and homozygous B-hCD3E/hCD20 mice (H/H), and analyzed by flow cytometry with anti-mouse CD3E antibody (Biolegend, 100321) and anti-human CD3E antibody (BD Pharmingen, 562426). Mouse CD3E was detectable in WT mice (+/+). Human CD3E was exclusively detectable in homozygous B-hCD3E/hCD20 mice (H/H) but not in WT mice (+/+).

Strain specific CD20 expression analysis in homozygous B-hCD3E/hCD20 mice by flow cytometry.
Splenocytes were collected from wild type (WT) mice (+/+) and homozygous B-hCD3E/hCD20 mice (H/H), and analyzed by flow cytometry with anti-mouse CD20 antibody (Biolegend, 152107) and anti-human CD20 antibody (Biolegend, 302305). Mouse CD20 was detectable in WT mice (+/+). Human CD20 was exclusively detectable in homozygous B-hCD3E/hCD20 mice (H/H) but not in WT mice (+/+).

Analysis of spleen leukocyte subpopulations by FACS.
Splenocytes were isolated from female C57BL/6 and B-hCD3E/hCD20 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 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-hCD3E/hCD20 mice were similar to those in the C57BL/6 mice, demonstrating that introduction of hCD3E/hCD20 in place of its mouse counterpart does not change the overall development, differentiation or distribution of these cell types in spleen.

Analysis of spleen T cell subpopulations by FACS.
Splenocytes were isolated from female C57BL/6 and B-hCD3E/hCD20 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 CD3 T cell population and used for further analysis as indicated here. B. Results of FACS analysis. Percent of CD8+ T cells, CD4+ T cells, and Treg cells in homozygous B-hCD3E/hCD20 mice were similar to those in the C57BL/6 mice, demonstrating that introduction of hCD3E/hCD20 in place of its mouse counterpart does not change the overall development, differentiation or distribution of these T cell sub types in spleen. Values are expressed as mean ± SEM.

Analysis of lymph node leukocyte subpopulations by FACS.
Leukocytes were isolated from female C57BL/6 and B-hCD3E/hCD20 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-hCD3E/hCD20 mice were similar to those in the C57BL/6 mice, demonstrating that introduction of hCD3E/hCD20 in place of its mouse counterpart does not change the overall development, differentiation or distribution of these cell types in lymph node.

Analysis of lymph node T cell subpopulations by FACS.
Leukocytes were isolated from female C57BL/6 and B-hCD3E/hCD20 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 CD3 T cell population and used for further analysis as indicated here. B. Results of FACS analysis. Percent of CD8+ T cells, CD4+ T cells, and Treg cells in homozygous B-hCD3E/hCD20 mice were similar to those in the C57BL/6 mice, demonstrating that introduction of hCD3E/hCD20 in place of its mouse counterpart does not change the overall development, differentiation or distribution of these T cell sub types in lymph node. Values are expressed as mean ± SEM.

Analysis of blood leukocyte subpopulations by FACS.
Blood cells were isolated from female C57BL/6 and B-hCD3E/hCD20 mice (n=3, 6 week-old). Flow cytometry analysis of the blood 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-hCD3E/hCD20 mice were similar to those in the C57BL/6 mice, demonstrating that introduction of hCD3E/hCD20 in place of its mouse counterpart does not change the overall development, differentiation or distribution of these cell types in blood.

Analysis of blood T cell subpopulations by FACS.
Blood cells were isolated from female C57BL/6 and B-hCD3E/hCD20 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 CD3 T cell population and used for further analysis as indicated here. B. Results of FACS analysis. Percent of CD8+ T cells, CD4+ T cells, and Treg cells in homozygous B-hCD3E/hCD20 mice were similar to those in the C57BL/6 mice, demonstrating that introduction of hCD3E/hCD20 in place of its mouse counterpart does not change the overall development, differentiation or distribution of these T cell sub types in blood. Values are expressed as mean ± SEM.

Analysis of thymus T cell subpopulations by FACS.
Thymocytes were isolated from female C57BL/6 and B-hCD3E/hCD20 mice (n=3, 6 week-old). Flow cytometry analysis of the thymocytes was performed to assess leukocyte subpopulations. A. Representative FACS plots. Single live CD45+ cells were gated for CD3 T cell population and used for further analysis as indicated here. B. Results of FACS analysis. Percent of CD8+ T cells, CD4+ T cells, and Treg cells in homozygous B-hCD3E/hCD20 mice were similar to those in the C57BL/6 mice, demonstrating that introduction of hCD3E/hCD20 in place of its mouse counterpart does not change the overall development, differentiation or distribution of these T cell sub types in thymus. Values are expressed as mean ± SEM. DN: double negative. DP: double positive.

The antibodies used here is provided by the client. Data was shown as Mean±SEM.