Basic Information
Description
The mouse B7h3 gene was replaced by human B7H3 coding sequence in B-hB7-H3 EL4 cells. Human B7-H3 is highly expressed on the surface of B-hB7-H3 EL4 cells.
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Targeting strategy
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Gene targeting strategy for B-hB7-H3 EL4 cells. The exogenous promoter and human B7H3 coding sequence was inserted to replace part of murine exon 3 and all of exon 4. The insertion disrupts the endogenous murine B7h3 gene, resulting in a non-functional transcript.
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Protein expression analysis
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B7-H3 expression analysis in B-hB7-H3 EL4 cells by flow cytometry. Single cell suspensions from wild-type EL4 and B-hB7-H3 EL4 cultures were stained with species-specific anti-B7-H3 antibody. Human B7-H3 was detected on the surface of B-hB7-H3 EL4 cells but not wild-type EL4 cells. The 1-H02 clone of B-hB7-H3 EL4 cells was used for in vivo experiments.
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Tumor growth curve & Body weight changes
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Subcutaneous homograft tumor growth of B-hB7-H3 EL4 cells. B-hB7-H3 EL4 cells (2×105) and wild-type EL4 cells (2×105) were subcutaneously implanted into C57BL/6N mice (female, 6-9-week-old, n=5). Tumor volume and body weight were measured twice a week. (A) Average tumor volume ± SEM. (B) Body weight (Mean± SEM). Volume was expressed in mm3 using the formula: V=0.5 X long diameter X short diameter2. As shown in panel A, B-hB7-H3 EL4 cells were able to establish tumors in vivo and can be used for efficacy studies.
Animals with tumor volume over 3000mm3 in the control group were euthanized.
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Tumor volume measurements
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Protein expression analysis of tumor cells
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B-hB7-H3 EL4 cells were subcutaneously transplanted into C57BL/6N mice (n=5), and on 27 days post inoculation, tumor cells were harvested and assessed for human B7-H3 expression by flow cytometry. As shown, human B7-H3 was highly expressed on the surface of tumor cells. Therefore, B-hB7-H3 EL4 cells can be used for in vivo efficacy studies of novel B7-H3 therapeutics.
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Tumor growth curve & Body weight changes
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Subcutaneous homograft tumor growth of B-hB7-H3 EL4 cells. B-hB7-H3 EL4 cells and wild-type MC38 cells were subcutaneously implanted into B-h4-1BB mice (female, 7-week-old, n=6). Tumor volume and body weight were measured twice a week. (A) Average tumor volume. (B) Body weight. Volume was expressed in mm3 using the formula: V=0.5 X long diameter X short diameter2. As shown in panel A, B-hB7-H3 EL4 cells were able to establish tumors in vivo and can be used for efficacy studies. Values are expressed as mean ± SEM.
