使用海马XF96分析仪评估钙刺激的线粒体生物能量学。

Assessing Calcium-Stimulated Mitochondrial Bioenergetics Using the Seahorse XF96 Analyzer.

作者信息

Wettmarshausen Jennifer, Perocchi Fabiana

机构信息

Institute for Diabetes and Obesity, Helmholtz Diabetes Center (HDC), Helmholtz Zentrum München and German National Diabetes Center (DZD), Neuherberg, Germany.

Department of Biochemistry, Gene Center Munich, Ludwig-Maximilians Universität München, Munich, Germany.

出版信息

Methods Mol Biol. 2019;1925:197-222. doi: 10.1007/978-1-4939-9018-4_18.

DOI:10.1007/978-1-4939-9018-4_18

PMID:30674029

Abstract

The development of fluorescence-based oxygen sensors coupled with microplate-based assays for quantitative bioenergetics analyses enables screening multiple experimental conditions at once with small biological material and in a timely manner. In this chapter, we outline detailed protocols and practical tips to design and perform controlled measurements of (a) respiratory and glycolytic metabolism of intact cells, (b) substrate-dependent respiration in permeabilized cells and isolated mitochondria, and (c) calcium-dependent regulation of mitochondrial bioenergetics with Seahorse XF Flux Analyzers.

摘要

基于荧光的氧传感器与基于微孔板的分析方法相结合用于定量生物能量学分析，使得能够使用少量生物材料同时及时筛选多种实验条件。在本章中，我们概述了详细的方案和实用技巧，以使用海马XF通量分析仪设计并进行以下各项的对照测量：(a) 完整细胞的呼吸和糖酵解代谢；(b) 透化细胞和分离线粒体中底物依赖性呼吸；以及 (c) 线粒体生物能量学的钙依赖性调节。