Linked Life Data

18340643

Source:http://linkedlifedata.com/resource/pubmed/id/18340643

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
5
pubmed:dateCreated
2008-4-23
pubmed:abstractText
Monitoring protein function with high throughput at individual cell level is of high interest both for basic research and diagnostic applications. For this, following the changes in fluorescence resonance energy transfer (FRET) between a donor/acceptor pair, genetically encoded in the proteins of interest, is a frequently used tool. As proteins attached to or located in the plasma membrane represent a considerable fraction of total proteins, there is a need for high throughput imaging techniques suited for observation of proteins in the cell membrane only. A system is presented, which allows rapid imaging of large areas via total internal reflection fluorescence microscopy (TIRFM) conditions, using a focus-hold system, multiwavelength excitation and dual color detection. The developed imaging system enables screening of large numbers of cells under TIRFM illumination combined with FRET imaging, thereby providing the means to record, e.g., FRET-efficiency of a membrane-associated protein labeled with a donor-acceptor pair. The capability of the system to perform live-FRET scanning with TIRFM on stoichiometric FRET constructs, reaching throughput of up to 1,000 cells/s at the optical resolution limit is demonstrated. A comparison with confocal microscopy shows that TIRFM offers a 4.2-fold advantage in our conditions over confocal microscopy in detecting contributions from membrane-localized proteins.
pubmed:commentsCorrections
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
May
pubmed:issn
1552-4930
pubmed:author
pubmed:copyrightInfo
(c) 2008 International Society for Advancement of Cytometry.
pubmed:issnType
Electronic
pubmed:volume
73
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
442-50
pubmed:meshHeading
pubmed:year
2008
pubmed:articleTitle
High throughput FRET screening of the plasma membrane based on TIRFM.
pubmed:affiliation
Center for Biomedical Nanotechnology, Upper Austrian Research GmbH, Linz, Austria.
pubmed:publicationType
Journal Article, Comparative Study, Research Support, Non-U.S. Gov't