Gene targeting strategy for B-hCD56 MC38 cells. The exogenous promoter and human CD56 coding sequence was inserted to replace part of murine exon 3 and all of exons 4-6. The insertion disrupts the endogenous murine Ncam1 gene, resulting in a non-functional transcript.

CD56 expression analysis in B-hCD56 MC38 cells by flow cytometry. Single cell suspensions from wild-type MC38 and B-hCD56 MC38 cultures were stained with species-specific anti-CD56 antibody. Human CD56 was detected on the surface of B-hCD56 MC38 cells but not wild-type MC38 cells. The 3-B10 clone of B-hCD56 MC38 cells was used for in vivo experiments.

Subcutaneous homograft tumor growth of B-hALB MC38 cells. B-hALB MC38 cells (1x106) and wild-type MC38 cells (5x105) were subcutaneously implanted into C57BL/6N mice (female, 6-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-hALB MC38 cells were able to establish tumors in vivo and can be used for efficacy studies.