A1 Vertaisarvioitu alkuperäisartikkeli tieteellisessä lehdessä
Liposome-based homogeneous luminescence resonance energy transfer
Tekijät: Pihlasalo S., Hara M., Hänninen P., Slotte J., Peltonen J., Härmä H.
Julkaisuvuosi: 2009
Lehti:: Analytical Biochemistry
Tietokannassa oleva lehden nimi: Analytical Biochemistry
Vuosikerta: 384
Numero: 2
Aloitussivu: 231
Lopetussivu: 237
Sivujen määrä: 7
ISSN: 0003-2697
DOI: https://doi.org/10.1016/j.ab.2008.09.035
Verkko-osoite: http://api.elsevier.com/content/abstract/scopus_id:56749173561
Tiivistelmä
There is an increasing need for developing simple assay formats for biomedical screening purposes. Assays on cell membranes have become important in studies of receptor-ligand interactions and signal pathways. Here luminescence energy transfer was studied on liposomes containing europium ion chelated to 4,4,4-trifluoro-1-(2-naphthalenyl)-1,3-butanedione and trioctylphosphine oxide. Energy transfer efficiency was characterized with biotin-streptavidin interaction, and a model assay concept for a homogeneous time-resolved luminescence resonance energy transfer (LRET) assay was developed. Acceptor-labeled streptavidin was bound to biotinylated lipids on the liposomes, leading to close proximity of the LRET pair. The liposome-based LRET assay was optimized for dye incorporation and concentration, biotinylation degree, liposome size, and kinetics. Sensitivity for a competitive biotin assay was at a picomolar range with a coefficient of variation from 7 to 20%. The developed lipid membrane-based system was feasible in separation free LRET assay concept with high sensitivity, indicating that the assay principle can potentially be used for biologically more relevant target molecules. © 2008 Elsevier Inc. All rights reserved.
There is an increasing need for developing simple assay formats for biomedical screening purposes. Assays on cell membranes have become important in studies of receptor-ligand interactions and signal pathways. Here luminescence energy transfer was studied on liposomes containing europium ion chelated to 4,4,4-trifluoro-1-(2-naphthalenyl)-1,3-butanedione and trioctylphosphine oxide. Energy transfer efficiency was characterized with biotin-streptavidin interaction, and a model assay concept for a homogeneous time-resolved luminescence resonance energy transfer (LRET) assay was developed. Acceptor-labeled streptavidin was bound to biotinylated lipids on the liposomes, leading to close proximity of the LRET pair. The liposome-based LRET assay was optimized for dye incorporation and concentration, biotinylation degree, liposome size, and kinetics. Sensitivity for a competitive biotin assay was at a picomolar range with a coefficient of variation from 7 to 20%. The developed lipid membrane-based system was feasible in separation free LRET assay concept with high sensitivity, indicating that the assay principle can potentially be used for biologically more relevant target molecules. © 2008 Elsevier Inc. All rights reserved.
Turun yliopiston Tutkimustietojärjestelmän portaali