Basic Information
Description
The mouse Epcam gene was replaced by the human EPCAM coding sequence in B-hEPCAM LLC1 cells. Human EPCAM is highly expressed on the surface of B-hEPCAM LLC1 cells.
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Gene targeting strategy
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Gene targeting strategy for B-hEPCAM LLC1 cells. The exogenous promoter and human EPCAM coding sequence was inserted to replace part of murine exon 4 and all of exons 5~7. The insertion disrupts the endogenous murine Epcam gene, resulting in a non-functional transcript.
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Protein expression analysis
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EPCAM protein expression analysis in B-hEPCAM LLC1 cells. Single cell suspensions from wild-type LLC1 and B-hEPCAM LLC1 cell cultures were stained with species-specific anti-EPCAM antibodies. Human EPCAM protein was detected on the surface of B-hEPCAM LLC1 cells but not wild-type LLC1 cells. The 1-B12 clone and the 1-F11 clone of B-hEPCAM LLC1 cells were used for in vivo experiments.
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Tumor growth curve in wild-type mice
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Subcutaneous homograft tumor growth of B-hEPCAM LLC1 cells. B-hEPCAM LLC1 cells (2×105) were subcutaneously implanted into C57BL/6 mice (female, 11-week-old, n=6). Tumor volume and body weight were measured three times 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-hEPCAM LLC1 cells were able to establish tumors in vivo, which can be used for efficacy studies.
Tumor volume and weight measurements
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Protein expression analysis post-inoculation
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B-hEPCAM LLC1 cells were subcutaneously transplanted into C57BL/6 mice (n=6), 22 days and 25 days post inoculation, tumor cells were harvested and assessed for human EPCAM protein expression by flow cytometry. As shown, human EPCAM protein was highly expressed on the surface of tumor cells. Therefore, B-hEPCAM LLC1 cells can be used for in vivo efficacy studies to test novel EPCAM therapeutics.
