当前位置: Nature » 细胞生物学 » 正文

Nat Protoc:华东理工大学杨弋教授开发新型细胞检测传感器

摘要 : 2016年6月30日,国际学术权威刊物自然出版集团旗下子刊《Nature Protocols》在线发表了华东理工大学药学院、生物反应器工程国家重点实验室特聘杨弋教授的一篇研究论文

 2016年6月30日,国际学术权威刊物自然出版集团旗下子刊《Nature Protocols》在线发表了华东理工大学药学院、生物反应器工程国家重点实验室特聘杨弋教授的一篇研究论文,论文报告称研究人员发明了对NAD+/NADH氧化还原状态高度敏感的一种传感器SoNar,该传感器可用于体内监测细胞的能量代谢。




In vivo monitoring of cellular energy metabolism using SoNar, a highly responsive sensor for NAD+/NADH redox state


NADH and its oxidized form NAD+ have a central role in energy metabolism, and their concentrations are often considered to be among the most important readouts of metabolic state. Here, we present a detailed protocol to image and monitor NAD+/NADH redox state in living cells and in vivo using a highly responsive, genetically encoded fluorescent sensor known as Sonar (sensor of NAD(H) redox). The chimeric Sonar protein was initially developed by inserting circularly permuted yellow fluorescent protein (cpYFP) into the NADH-binding domain of Rex protein fromThermus aquaticus (T-Rex). It functions by binding to either NAD+ or NADH, thus inducing protein conformational changes that affect its fluorescent properties. We first describe steps for how to establish SoNar-expressing cells, and then discuss how to use the system to quantify the intracellular redox state. This approach is sensitive, accurate, simple and able to report subtle perturbations of various pathways of energy metabolism in real time. We also detail the application of Sonar to high-throughput chemical screening of candidate compounds targeting cell metabolism in a microplate-reader-based assay, along with in vivo fluorescence imaging of tumor xenografts expressing Sonar in mice. Typically, the approximate time frame for fluorescence imaging of Sonar is 30 min for living cells and 60 min for living mice. For high-throughput chemical screening in a 384-well-plate assay, the whole procedure generally takes no longer than 60 min to assess the effects of 380 compounds on cell metabolism.

来源: Nature Protocols 浏览次数:0


RSS订阅 - 填写您的邮件地址,订阅我们的精彩内容: - 网站地图
网站联系电话:020-87540820 备案号:粤ICP备11050685号-8 增值电信业务经营许可证:粤B2-20120479
©2011-2015 生物帮 All rights reserved.