Gaikwad Sadanand M, Heneka Michael T
University of Bonn, Bonn, Germany.
Methods Mol Biol. 2013;1041:185-97. doi: 10.1007/978-1-62703-520-0_18.
Microglial cell function receives increasing interest. To date, the majority of experiments are performed by using immortalized microglia-like cells or primary microglia prepared from pre- or postnatal rodent brain. As those may not adequately reflect the microglial biology in the adult brain, this protocol advocates a procedure which allows for the isolation, purification, and subsequent analysis of microglial cells. Once isolated, the principal state of activation, M1 or M2, can be determined in adult microglia using fluorescence-activated cell sorting, quantitative PCR, and/or Western blotting. Likewise, adult microglia generated by this protocol can be used for functional analysis through cell cultivation for a limited time.
小胶质细胞功能越来越受到关注。迄今为止，大多数实验是使用永生化的小胶质细胞样细胞或从出生前或出生后的啮齿动物大脑中制备的原代小胶质细胞进行的。由于这些细胞可能无法充分反映成年大脑中的小胶质细胞生物学特性，本方案推荐一种能够分离、纯化并随后分析小胶质细胞的方法。一旦分离出来，就可以使用荧光激活细胞分选、定量PCR和/或蛋白质印迹法来确定成年小胶质细胞的主要激活状态，即M1或M2。同样，通过本方案产生的成年小胶质细胞可在有限时间内通过细胞培养用于功能分析。
