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  • Title: [Knockout serum replacement improves establishment efficiency of C57BL/6J mouse embryonic stem cell line].
    Author: Wang H, Tian HB, Chen J, Sha HY, Chen JQ, Cheng GX.
    Journal: Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi; 2007 Mar; 23(3):269-72. PubMed ID: 17343798.
    Abstract:
    AIM: To eliminate the influence of serum on self-renewal of embryonic stem cells (ESCs), knockout serum replacement (KSR), a defined formulation, was used to replace serum for the establishment of C57BL/6J mouse ESC line. METHODS: C57BL/6J mouse blastocysts collected at 3.5 days post coitum (d.p.c.) were cultured in the medium supplemented with KSR. In control experiment, KSR was substituted by fetal bovine serum (FBS). When ESC line was established, the morphology of ESCs, the expression of alkaline phosphatase and oct-4, and the karyotype and differentiating ability of ESCs were analyzed. RESULTS: 13 blastocysts were cultured in the medium supplemented with KSR and one ESC line (MES-1) was established with a normal and stable XX karyotype after cultured for more than 20 passages, and then the high expression of alkaline phosphatase and oct-4 was detected. When cultured in suspension, MES-1 formed embryoid bodies. When inoculated subcutaneously into nude mice, MES-1 formed teratoma. After injected into ICR mouse blastocysts collected at 3.5 d.p.c., MES-1 incorporated into the inner cell mass of the host blastocyst and contributed to the development of a chimera. In control experiment, no ESC lines were cultured for more than 3 passages. CONCLUSION: KSR can be efficiently used to isolate and culture C57BL/6J mouse ESCs, which can eliminate traditional prescreening of FBS suitable for isolation and culture of ESCs.
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