Identification of gap junction blockers using automated fluorescence microscopy imaging.
Gap junctions coordinate electrical signals and facilitate metabolic synchronization between cells. In this study, the authors have developed a novel assay for the identification of gap junction blockers using fluorescence microscopy imaging-based high-content screening technology. In the assay, the communication between neighboring cells through gap junctions was measured by following the redistribution of a fluorescent marker. The movement of calcein dye from dye-loaded donor cells to dye-free acceptor cells through gap junctions overexpressed on cell surface membranes was monitored using automated fluorescence microscopy imaging in a high-throughput compatible format. The fluorescence imaging technology consisted of automated focusing, image acquisition, image processing, and data mining. The authors have successfully performed a high-throughput screening of a 486,000- compound program with this assay, and they were able to identify false positives without additional experiments. Selective and pharmacologically interesting compounds were identified for further optimization.
Lead Discovery Technology, Lead Generation, Aventis Pharmaceutical, Bridgewater, NJ 08807, USA. julie.li@aventis.com
Zhuyin
Z
Li
Yongping
Y
Yan
Elaine A
EA
Powers
Xiaoyou
X
Ying
Khurram
K
Janjua
Tina
T
Garyantes
Bruce
B
Baron
D000818:Animals; D001331:Automation; D002450:Cell Communication; D002455:Cell Division / Q000502:physiology; D016543:Central Nervous System Neoplasms / Q000188:drug therapy / Q000473:pathology; D004305:Dose-Response Relationship, Drug; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D005189:False Positive Reactions; D005452:Fluoresceins / Q000378:metabolism; D005456:Fluorescent Dyes / Q000378:metabolism; D017629:Gap Junctions / Q000187:drug effects*; D005910:Glioma / Q000188:drug therapy / Q000473:pathology; D007091:Image Processing, Computer-Assisted / Q000379:methods*; D008469:Meclofenamic Acid / Q000494:pharmacology; D008856:Microscopy, Fluorescence / Q000379:methods*; D051381:Rats; D012680:Sensitivity and Specificity; D013997:Time Factors; D014407:Tumor Cells, Cultured
D005452:Fluoresceins; D005456:Fluorescent Dyes; D008469:Meclofenamic Acid; C007740:fluorexon