Isolation and short-term culture of primary keratinocytes, hair follicle populations and dermal cells from newborn mice and keratinocytes from adult mice for in vitro analysis and for grafting to immunodeficient mice
Isolation and short-term culture of primary keratinocytes, hair follicle populations and dermal cells from newborn mice and keratinocytes from adult mice for in vitro analysis and for grafting to immunodeficient mice
复制标题
DOI：
10.1038/nprot.2008.50
复制
发表时间：
2008-01-01
期刊：
影响因子：
14.8
通讯作者：
Yuspa, Stuart H.
中科院分区：
文献类型：
--
作者：
Lichti, Ulrike;Anders, Joanna;Yuspa, Stuart H.
从新生和成年小鼠表皮制备和培养原代角质形成细胞的方案在过去的35年中不断发展。该方案现在常规应用于各种遗传背景的小鼠，用于分化和细胞转化中的信号传导途径的体外研究，以及用于评估免疫缺陷小鼠上的细胞移植物中改变的角质形成细胞的体内表型。在该方法的开发和应用中至关重要的是观察到角质形成细胞在低钙浓度的培养基中增殖，但在初始附着期后在钙浓度> 0.07 mM时迅速分化。从10只新生小鼠制备原代角质形成细胞需要2-3小时的动手时间。还提供了相关的程序：制备未成熟的毛囊芽，从新生小鼠发育真皮毛囊和成纤维细胞，从成年小鼠制备原代角质形成细胞和将细胞混合物移植到无胸腺裸鼠上。
Protocols for preparing and culturing primary keratinocytes from newborn and adult mouse epidermis have evolved over the past 35 years. This protocol is now routinely applied to mice of various genetic backgrounds for in vitro studies of signaling pathways in differentiation and cell transformation, and for assessing the in vivo phenotype of altered keratinocytes in grafts of cells on immunodeficient mice. Crucial in the development and application of the procedure was the observation that keratinocytes proliferate in media of low calcium concentration, but rapidly commit to differentiation at calcium concentrations > 0.07 mM after the initial attachment period. Preparing primary keratinocytes from ten newborn mice requires 2-3 h of hands-on time. Related procedures are also provided: preparing immature hair follicle buds, developing dermal hair follicles and fibroblasts from newborn mice, preparing primary keratinocytes from adult mice and grafting cell mixtures on athymic nude mice.