Linked Life Data

18931453

Source:http://linkedlifedata.com/resource/pubmed/id/18931453

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
1
pubmed:dateCreated
2008-10-20
pubmed:abstractText
Fluorescence cross-correlation spectroscopy (FCCS) is used as a powerful technique to analyze molecular interactions both in vitro and in vivo. This method basically requires two laser excitations for two target molecules labeled with fluorophores of different colors. Their coincidence in a microscopic detection volume is analyzed using two detectors. Any overlap of emission spectra of the two fluorophores, however, gives rise to false-positive data about their interaction. To overcome this problem, we have developed a new FCCS system, in which two excitation lasers are switched alternately by modulation using an acousto-optic tunable filter (AOTF). In this report, we demonstrate the feasibility of switching FCCS for enzymatic cleavage of proteins in living cells. A fusion protein of two fluorophores (EGFP and mRFP) with a cleavage site of caspase-3 inserted was expressed in HeLa cells, and proteolysis assay was performed during apoptotic cell death. Due to the absence of cross-talk signals, the FCCS measurement with the switching function gave a large change in relative cross-correlation amplitude after protein cleavage. Hence, switching FCCS enables more reliable measurement of molecular interactions than conventional FCCS.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:issn
1347-3700
pubmed:author
pubmed:issnType
Electronic
pubmed:volume
33
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
143-50
pubmed:meshHeading
pubmed:year
2008
pubmed:articleTitle
Cross-talk-free fluorescence cross-correlation spectroscopy by the switching method.
pubmed:affiliation
Olympus Corporation, Tokyo, Japan. yas_takahashi@ot.olympus.co.jp
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't, Evaluation Studies