Athanasiadis Anastasios, Ambele Melvin A, Pepper Michael S
Institute for Cellular and Molecular Medicine, Department of Medical Immunology, and SAMRC Extramural Unit for Stem Cell Research and Therapy, Faculty of Health Sciences, University of Pretoria, Pretoria, South Africa.
Methods Mol Biol. 2025;2938:125-131. doi: 10.1007/978-1-0716-4607-6_15.
Adipose-derived mesenchymal stromal/stem cells (AD-MSCs) offer a promising cell source for regenerative medicine due to their easy accessibility and potential application across a wide range of disorders. This chapter describes a standardized enzymatic isolation protocol to extract AD-MSCs from both solid adipose tissue and lipoaspirate. Adipose tissue samples are initially washed with phosphate-buffered saline (PBS) to remove blood contaminants. Tissue is then digested enzymatically with type I collagenase under controlled temperature and agitation to release the stromal vascular fraction (SVF) containing AD-MSCs. The enzyme activity is neutralized using complete growth medium followed by centrifugation to separate the cell pellet from unwanted material. The resulting cell pellet is resuspended and filtered to eliminate residual tissue fragments. After seeding the cells in culture, AD-MSCs in the SVF are characterized by adherence to plastic and the expression of specific cell surface marker (e.g., CD73, CD90, CD105), confirming their identity. This protocol provides a method for isolating viable AD-MSCs for downstream applications.
脂肪来源的间充质基质/干细胞（AD-MSCs）因其易于获取且在多种疾病中具有潜在应用价值，为再生医学提供了一种有前景的细胞来源。本章介绍一种标准化的酶分离方案，用于从固体脂肪组织和抽脂物中提取AD-MSCs。脂肪组织样本首先用磷酸盐缓冲盐水（PBS）洗涤，以去除血液污染物。然后在控制温度和搅拌的条件下，用I型胶原酶对组织进行酶解，以释放包含AD-MSCs的基质血管成分（SVF）。使用完全生长培养基中和酶活性，随后进行离心，以将细胞沉淀与不需要的物质分离。将所得细胞沉淀重悬并过滤，以去除残留的组织碎片。将细胞接种到培养物中后，SVF中的AD-MSCs通过贴壁于塑料培养皿以及特定细胞表面标志物（如CD73、CD90、CD105）的表达来进行鉴定，从而确认其身份。该方案提供了一种分离有活力的AD-MSCs用于下游应用的方法。
