Tehrani Kayvan Forouhesh, Pendleton Emily G, Southern William M, Call Jarrod A, Mortensen Luke J
Regenerative Bioscience Center, Rhodes Center for ADS, University of Georgia, Athens, GA 30602, USA.
Department of Kinesiology, University of Georgia, Athens, GA 30602, USA.
Biomed Opt Express. 2017 Dec 19;9(1):254-259. doi: 10.1364/BOE.9.000254. eCollection 2018 Jan 1.
Cell metabolism and viability are directly reflected in their mitochondria. Imaging-based analysis of mitochondrial morphological structure, size and dynamic characteristics can therefore provide critical insight into cell function. However, mitochondria are often very abundant, and due to their close to diffraction-limit size, it is often non-trivial to distinguish a tubular or large mitochondrion from an ensemble of punctate mitochondria. In this paper, we use membrane potential dependent fluorescence fluctuations of individual mitochondria to resolve them using an approach similar to single molecule localization microscopy. We use 2-photon microscopy to image mitochondrial intensity fluctuations at 200 μm deep inside an intact in-vivo mouse soleus muscle. By analyzing the acquired images, we can reconstruct images with an extra layer of information about individual mitochondria, separated from their ensemble. Our analysis shows a factor of 14 improvement in detection of mitochondria.
细胞代谢和活力直接反映在其线粒体中。因此，基于成像的线粒体形态结构、大小和动态特征分析能够为细胞功能提供关键见解。然而，线粒体通常数量众多，并且由于其大小接近衍射极限，从点状线粒体集合中区分出管状或大型线粒体往往并非易事。在本文中，我们利用单个线粒体的膜电位依赖性荧光波动，采用类似于单分子定位显微镜的方法来分辨它们。我们使用双光子显微镜对完整的体内小鼠比目鱼肌内200μm深处的线粒体强度波动进行成像。通过分析获取的图像，我们可以重建包含关于单个线粒体的额外信息层的图像，这些单个线粒体与其集合体分离。我们的分析表明线粒体检测能力提高了14倍。
