Authors

Aoki, K., & Matsuda, M.

Affiliations

Laboratory of Bio-imaging and Cell Signalling, Graduate School of Bio-studies, Kyoto University, Kyoto, Japan. Department of Pathology and Biology of Diseases, Graduate School of Medicine, Kyoto University, Kyoto, Japan.

Application Area

Science Technology

FRET

Abstract

Protein kinases are key modulators of intracellular signal transduction cascades, which determine various events in neuronal cells such as replication and differentiation. For many years, protein kinases were analysed mostly by biochemical methods, which could handle the cells only en masse. For a better under- standing of the role of kinases in neuronal cells, one would like to know the subcellular distribution of kinase activities and to follow a particular kinase activity for a specific period in a single cell. Genetically encoded biosensors based on the principle of Förster (or fluorescence) resonance energy transfer (FRET) and fluorescent proteins have been developed to accommodate such requirements. The method involves expression of the FRET biosensors in neuronal cells, time-lapse imaging under fluorescence microscopes, image processing, and quantification of FRET. This technique could be applicable to living organisms ranging from Caenorhabditis elegans to mouse, permitting visualization of spatio-temporal regulation of kinase activities and systemic understanding of the signalling networks in living animals.

Extract

… “– LED illumination system, CoolLED precisExcite (Molecular device)”…

Product Associated Features

pE-100: A range of compact, simple-to-use, single wavelength illumination systems for screening fluorescence.

Diascopic Technique

Bright-Field

Live Cell Issues

Photobleaching

Product Type

pE-100

Journal

Nature protocols

Year of Publication

2009

Country of Publication

Japan