Investigating autophagy: quantitative morphometric analysis using electron microscopy.
Investigating autophagy: quantitative morphometric analysis using electron microscopy.
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DOI：
10.4161/auto.6.2.10439
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发表时间：
2010-02
期刊：
影响因子：
13.3
通讯作者：
Oberley TD
中科院分区：
文献类型：
--
作者：
Swanlund JM;Kregel KC;Oberley TD
自噬是真核细胞内细胞质和细胞器的一种代偿性降解途径。自噬过程可以通过以选择性或非选择性的方式去除受损的蛋白质和细胞器来提供“管家”功能，以便在应激后发挥保护作用。值得注意的是，在发现自噬更像是一个靶向过程而不是随机过程之后，自噬的作用与许多正常细胞和疾病过程有关。有几种方法被常规地用于监测整个自噬过程。微管相关蛋白轻链3是酵母Atg 8的哺乳动物同源物，已被广泛用作监测许多细胞类型中自噬的特异性标志物。虽然监测自噬通量非常重要，但通过电子显微镜（EM）进行详细分析以评估各种自噬结构的变化，量化所涉及的区域，并确定是否有任何特定的细胞器或细胞质区域被靶向降解也是有益的。下面的文章描述了使用透射电镜定位和定量自噬亚细胞区域的方法。还讨论了采用免疫金EM的特定蛋白质的亚细胞定位的方法;这种方法变得特别有用，在检测细胞内稳态的早期变化，可能会发生在后来的迹象细胞损伤可以观察到形态学。
Autophagy is a compensatory pathway involving isolation and subsequent degradation of cytosolic material and organelles in eukaryotic cells. The autophagic process can provide a “housekeeping” function by removing damaged proteins and organelles in a selective or nonselective fashion in order to exert a protective effect following stress. Remarkably, after being discovered to be much more of a targeted process than a random one, the role of autophagy became implicated in many normal cellular and disease processes. Several methodologies are routinely employed to monitor the entire autophagic process. Microtubule-associated protein light chain 3, a mammalian homolog of yeast Atg8, has been widely used as a specific marker to monitor autophagy in numerous cell types. While monitoring autophagic flux is extremely important, it is also beneficial to perform a detailed analysis by electron microscopy (EM) to evaluate changes in various autophagic structures, quantify the areas involved, and determine if any particular organelle(s) or area of the cell cytoplasm is being targeted for degradation. The following article describes methods to localize and quantify subcellular areas of autophagy using transmission EM. Also discussed are methods for subcellular localization of specific proteins by employing immunogold EM; this method becomes particularly useful in detecting early changes in cellular homeostasis that may occur before later signs of cellular insult can be observed morphologically.
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影响因子：
2
作者：
Gundersen, HJG;Jensen, EBV;Nielsen, J
通讯作者：
Nielsen, J
影响因子：
78.5
作者：
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影响因子：
2.9
作者：
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通讯作者：
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影响因子：
2
作者：
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通讯作者：
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影响因子：
3.2
作者：
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通讯作者：
Oberley, Terry D.