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Quantitative detection of cell surface protein expression by time-resolved fluorimetry

Huttunen, Roope J.; O'Riordan, Tomas C.; Härkönen, Pirkko LU ; Soini, Juhani T.; Meltola, Niko J.; Hanninen, Pekka E. and Soini, Aleksi E. (2007) In Luminescence 22(3). p.163-170
Abstract
A method is introduced for quantitative detection of cell surface protein expression. The method is based on immunocytochemistry, the use of long decay time europium(III) chelate and platinum(II) porphyrin labels, and detection of photoluminescence emission from adhered cells by time-resolved fluorimetry. After immunocytochemistry, the assay wells are evaporated to dryness and measured in the dry state. This protocol allows repeated and postponed analysis and microscopy imaging. In order to investigate the performance of the method, we chose expression of intercellular adhesion molecule-1 (ICAM-1) of endothelial cell line EAhy926 as a research target. The expression of ICAM-1 on the cells was enhanced by introduction of a cytokine, tumour... (More)
A method is introduced for quantitative detection of cell surface protein expression. The method is based on immunocytochemistry, the use of long decay time europium(III) chelate and platinum(II) porphyrin labels, and detection of photoluminescence emission from adhered cells by time-resolved fluorimetry. After immunocytochemistry, the assay wells are evaporated to dryness and measured in the dry state. This protocol allows repeated and postponed analysis and microscopy imaging. In order to investigate the performance of the method, we chose expression of intercellular adhesion molecule-1 (ICAM-1) of endothelial cell line EAhy926 as a research target. The expression of ICAM-1 on the cells was enhanced by introduction of a cytokine, tumour necrosis factor-alpha (TNF alpha). The method gave signal: background ratios (S:B) of 20 and 9 for europium and platinum labels, respectively, whereas prompt fluorescent FITC label gave a S:13 of 3. Screening window coefficients (=Z'-factor) were >0.5 for all the three labels, thus indicating a score for an excellent screening assay. In conclusion, the method appears to be an appropriate choice for protein expression analysis, both in high-throughput screening applications, and for detailed sample investigation by fluorescent microscopy imaging. Copyright (C) 2007 John Wiley & Sons, Ltd. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
keywords
endothelial cells, ICAM-1, time-resolved fluorescence, immunocytochemistry, protein expression analysis, drug discovery
in
Luminescence
volume
22
issue
3
pages
163 - 170
publisher
John Wiley & Sons
external identifiers
  • wos:000247841000002
  • scopus:34347272700
ISSN
1522-7243
DOI
10.1002/bio.943
language
English
LU publication?
yes
id
80ace16b-333e-43b7-a126-19440452e94e (old id 646308)
date added to LUP
2007-12-12 10:04:00
date last changed
2017-01-01 04:44:03
@article{80ace16b-333e-43b7-a126-19440452e94e,
  abstract     = {A method is introduced for quantitative detection of cell surface protein expression. The method is based on immunocytochemistry, the use of long decay time europium(III) chelate and platinum(II) porphyrin labels, and detection of photoluminescence emission from adhered cells by time-resolved fluorimetry. After immunocytochemistry, the assay wells are evaporated to dryness and measured in the dry state. This protocol allows repeated and postponed analysis and microscopy imaging. In order to investigate the performance of the method, we chose expression of intercellular adhesion molecule-1 (ICAM-1) of endothelial cell line EAhy926 as a research target. The expression of ICAM-1 on the cells was enhanced by introduction of a cytokine, tumour necrosis factor-alpha (TNF alpha). The method gave signal: background ratios (S:B) of 20 and 9 for europium and platinum labels, respectively, whereas prompt fluorescent FITC label gave a S:13 of 3. Screening window coefficients (=Z'-factor) were >0.5 for all the three labels, thus indicating a score for an excellent screening assay. In conclusion, the method appears to be an appropriate choice for protein expression analysis, both in high-throughput screening applications, and for detailed sample investigation by fluorescent microscopy imaging. Copyright (C) 2007 John Wiley & Sons, Ltd.},
  author       = {Huttunen, Roope J. and O'Riordan, Tomas C. and Härkönen, Pirkko and Soini, Juhani T. and Meltola, Niko J. and Hanninen, Pekka E. and Soini, Aleksi E.},
  issn         = {1522-7243},
  keyword      = {endothelial cells,ICAM-1,time-resolved fluorescence,immunocytochemistry,protein expression analysis,drug discovery},
  language     = {eng},
  number       = {3},
  pages        = {163--170},
  publisher    = {John Wiley & Sons},
  series       = {Luminescence},
  title        = {Quantitative detection of cell surface protein expression by time-resolved fluorimetry},
  url          = {http://dx.doi.org/10.1002/bio.943},
  volume       = {22},
  year         = {2007},
}