Demonstration of a visual cell-based assay for screening glucose transporter 4 translocation modulators in real time

Journal of biosciences

Maleppillil Vavachan Vijayakumar¹  Amrendra Kumar Ajay¹  Manoj Kumar Bhat1¹ 

[1] National Centre for Cell Science, NCCS Complex, Ganeshkhind, Pune 411 007, India$$

关键词: CHO cells;    eGFP;    GLUT4;    live cell imaging;    natural product;    qualitative assay;    translocation;   

DOI  :  

来源: Indian Academy of Sciences

【 摘 要 】

Insulin-stimulated translocation of glucose transporter 4 (GLUT4) to cell membrane leading to glucose uptake is the rate-limiting step in diabetes. It is also a defined target of antidiabetic drug research. Existing GLUT4 translocation assays are based on time-consuming immunoassays and are hampered by assay variability and low sensitivity. We describe a real-time, visual, cell-based qualitative GLUT4 translocation assay using CHO-HIRc-myc-GLUT4eGFP cells that stably express myc- and eGFP-tagged GLUT4 in addition to human insulin receptor (HIRc). GLUT4 translocation is visualized by live cell imaging based on GFP fluorescence by employing a cooled charge-coupled device camera attached to a fluorescent microscope. This video imaging method and further quantitative analysis of GLUT4 on the cell membrane provide rapid and foolproof visual evidence that this method is suitable for screening GLUT4 translocation modulators.

【 授权许可】

Unknown