Nadri Samad, Yazdani Shahin
Medical Biotechnology and Medical Nanotechnology Department, Faculty of Medicine, Zanjan University of Medical Science, Zanjan, Iran.
Ophthalmic Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran.
Curr Protoc Stem Cell Biol. 2015 May 1;33:1F.14.1-8. doi: 10.1002/9780470151808.sc01f14s33.
Here we describe a simple protocol for the isolation and culture of mesenchymal stem cells (MSCs) from conjunctiva stromal tissue, with applications to stem cell biology and regenerative medicine. This protocol is based on an explant culture protocol for the adhesion and migration of MSCs from tissue biopsy in primary culture and expansion of the MSC population by passaging cells on the surface of plastic culture dishes. Conjunctiva mesenchymal stem cells (CJMSCs) are generally isolated from small biopsies of human conjunctival tissue. Epithelial sheets are carefully loosened and removed by enzyme treatment after 18 hr and placed in fresh medium supplemented with growth factors. When the cells migrating from tissue segments in primary cultures become nearly confluent, the culture is treated with trypsin. A purified population of CJMSCs that retain proliferation and differentiation potential for many passages can be obtained 2 weeks after the initiation of culture.
在此，我们描述了一种从结膜基质组织中分离和培养间充质干细胞（MSCs）的简单方案，该方案可应用于干细胞生物学和再生医学领域。此方案基于一种外植体培养方案，用于在原代培养中使MSCs从组织活检中黏附并迁移，并通过在塑料培养皿表面传代细胞来扩增MSCs群体。结膜间充质干细胞（CJMSCs）通常从人结膜组织的小活检样本中分离得到。18小时后，通过酶处理小心地松解并去除上皮片，然后将其置于添加了生长因子的新鲜培养基中。当原代培养中从组织片段迁移出的细胞接近汇合时，用胰蛋白酶处理培养物。在培养开始2周后，可获得一群纯化的CJMSCs，它们在许多传代中都保留增殖和分化潜能。
