Expression, purification and use of recombinant annexin V for the detection of apoptotic cells
Expression, purification and use of recombinant annexin V for the detection of apoptotic cells
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DOI：
10.1038/nprot.2009.143
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发表时间：
2009-01-01
期刊：
影响因子：
14.8
通讯作者：
Martin, Seamus J.
中科院分区：
文献类型：
--
作者：
Logue, Susan E.;Elgendy, Mohamed;Martin, Seamus J.
细胞凋亡是程序性细胞死亡的一种模式，广泛用于在发育、组织稳态、感染或响应损伤期间消除细胞。凋亡细胞质膜的改变触发吞噬细胞对这些细胞的识别和吞噬。使用荧光标记的膜联蛋白V测量质膜磷脂酰丝氨酸外化被广泛用于检测凋亡细胞。在这里，我们描述了重组膜联蛋白V的细菌表达，纯化和FITC标记的协议。通过遵循本协议中概述的方法，有可能在3天内产生毫克量的重组膜联蛋白V。我们还描述了通过流式细胞术或荧光显微镜评估膜联蛋白V与细胞群结合的方法。
Apoptosis is a mode of programmed cell death that is widely used to eliminate cells during development, tissue homeostasis, infection or in response to injury. Alterations to the plasma membranes of apoptotic cells trigger recognition and engulfment of such cells by phagocytes. Measurement of plasma membrane phosphatidylserine externalization, using fluorescently labeled annexin V, is widely used for the detection of apoptotic cells. Here we describe protocols for bacterial expression, purification and FITC labeling of recombinant annexin V. By following the method outlined in this protocol, it is possible to produce milligram amounts of recombinant annexin V within 3 d. We also describe a method for the assessment of annexin V binding to cell populations by flow cytometry or fluorescence microscopy.