Basic Information
Description
The mouse Lrrc15 gene was replaced by human LRRC15 coding sequence in B-hLRRC15 MC38 cells. Human LRRC15 is highly expressed on the surface of B-hLRRC15 MC38 cells. B-hLRRC15 MC38 cells have the capability to establish tumors in vivo and can be used for efficacy studies.
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Gene targeting strategy
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Gene targeting strategy for B-hLRRC15 MC38 cells. The exogenous promoter and human LRRC15 coding sequence was inserted to replace part of murine exon 2. The insertion disrupts the endogenous murine Lrrc15 gene, resulting in a non-functional transcript.
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Protein expression analysis
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LRRC15 expression analysis in B-hLRRC15 MC38 cells by flow cytometry. Single cell suspensions from wild-type MC38 and B-hLRRC15 MC38 cultures were stained with an anti-human LRRC15 antibody and analyzed by flow cytometry. Human LRRC15 was detected on the surface of B-hLRRC15 MC38 cells but not on wild-type MC38 cells. Clone 2-C04 was used for in vivo experiments.
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Tumor growth curve and body weight changes
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Subcutaneous homograft tumor growth of B-hLRRC15 MC38 cells. B-hLRRC15 MC38 cells (5×106) and wild-type MC38 cells (5×105) were subcutaneously implanted into C57BL/6 mice (female, 7-9-week-old, n=8). 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-hLRRC15 MC38 cells were able to establish tumors in vivo and can be used for efficacy studies.
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Protein expression analysis - post-inoculation
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B-hLRRC15 MC38 cells were subcutaneously transplanted into C57BL/6 mice (n=6), 28 days post-inoculation, tumor cells were harvested and assessed for human LRRC15 expression by flow cytometry. As shown, human LRRC15 was highly expressed on the surface of tumor cells. Therefore, B-hLRRC15 MC38 cells can be used for in vivo efficacy studies testing novel LRRC15 therapeutics.
