Bonekamp Nina A, Islinger Markus, Lázaro Maria Gómez, Schrader Michael
Center for Cell Biology and Department of Biology, University of Aveiro, Aveiro, Portugal.
Methods Mol Biol. 2013;931:467-82. doi: 10.1007/978-1-62703-056-4_24.
Peroxisomes and mitochondria are essential subcellular organelles in mammals. Interestingly, recent studies have elucidated that these highly dynamic and plastic organelles exhibit a much closer interrelationship than previously assumed. Peroxisomes and mitochondria are metabolically linked organelles, which are cooperating and cross-talking. They share key components of their division machinery and cooperate in antiviral signaling and defense. As peroxisomal alterations in metabolism, biogenesis, dynamics, and proliferation have the potential to influence mitochondrial morphology and functions (and vice versa), there is currently great interest in the detection of both organelles under different experimental conditions. Here, we present protocols used successfully in our laboratory for the dual detection of peroxisomes and mitochondria in cultured mammalian cells. We address double immunofluorescence and fluorescence-based techniques as well as reagents to investigate organelle dynamics, morphological alterations, and organelle-specific targeting of proteins. In addition, we describe the application of diaminobenzidine cytochemistry on cultured cells to specifically label peroxisomes in ultrastructural studies.
过氧化物酶体和线粒体是哺乳动物体内重要的亚细胞器。有趣的是，最近的研究表明，这些高度动态和可塑性的细胞器之间的相互关系比以前认为的要密切得多。过氧化物酶体和线粒体是代谢相关的细胞器，它们相互协作并相互交流。它们共享分裂机制的关键成分，并在抗病毒信号传导和防御中协同作用。由于过氧化物酶体在代谢、生物发生、动态变化和增殖方面的改变有可能影响线粒体的形态和功能（反之亦然），目前人们对在不同实验条件下检测这两种细胞器非常感兴趣。在这里，我们介绍了我们实验室成功用于在培养的哺乳动物细胞中双重检测过氧化物酶体和线粒体的方法。我们讨论了双重免疫荧光和基于荧光的技术以及用于研究细胞器动态变化、形态改变和蛋白质的细胞器特异性靶向的试剂。此外，我们描述了二氨基联苯胺细胞化学在培养细胞上的应用，以在超微结构研究中特异性标记过氧化物酶体。
