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Titolo:
Simultaneous analysis of the cyan, yellow and green fluorescent proteins by flow cytometry using single-laser excitation at 458 nm
Autore:
Beavis, AJ; Kalejta, RF;
Indirizzi:
Princeton Univ, Lewis Thomas Lab, Flow Cytometry Core Facil, Princeton, NJ08544 USA Princeton Univ Princeton NJ USA 08544 Core Facil, Princeton, NJ08544 USA Princeton Univ, Dept Biol Mol, Princeton, NJ 08544 USA Princeton Univ Princeton NJ USA 08544 t Biol Mol, Princeton, NJ 08544 USA
Titolo Testata:
CYTOMETRY
fascicolo: 1, volume: 37, anno: 1999,
pagine: 68 - 73
SICI:
0196-4763(19990901)37:1<68:SAOTCY>2.0.ZU;2-2
Fonte:
ISI
Lingua:
ENG
Soggetto:
CELLS; FLUOROCHROMES; MUTATIONS; EMISSION; MUTANTS; GFP;
Keywords:
green yellow cyan fluorescent proteins; EGFP; EYFP; ECFP; single laser excitation; flow cytometry; 458 nm excitation;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Life Sciences
Citazioni:
32
Recensione:
Indirizzi per estratti:
Indirizzo: Beavis, AJ Princeton Univ, Lewis Thomas Lab, Flow Cytometry Core Facil, Washington Rd, Princeton, NJ 08544 USA Princeton Univ Washington Rd PrincetonNJ USA 08544 J 08544 USA
Citazione:
A.J. Beavis e R.F. Kalejta, "Simultaneous analysis of the cyan, yellow and green fluorescent proteins by flow cytometry using single-laser excitation at 458 nm", CYTOMETRY, 37(1), 1999, pp. 68-73

Abstract

Background: Development of spectrally distinct green fluorescent protein (GFP) variants has allowed for simultaneous now cytometric detection of two different colored mutants expressed in a single cell. However, the dual-laser methods employed in such experiments are not widely applicable since they require a specific, expensive laser, and single-laser analysis at 488 nm exhibits considerable spectral overlap. The purpose of this work was to evaluate detection of enhanced cyan fluorescent protein (ECFP) in combination with the enhanced green (EGFP) and enhanced yellow (EYFP) fluorescent proteins by now cytometry. Methods: Cells transfected with expression constructs for EGFP, EYFP, or ECFP were analyzed by now cytometry using excitation wavelengths at 458, 488, or 514 nm. Fluorescence signals were separated with a custom optical filter configuration: 525 nm shortpass and 500 nm longpass dichroics; 480/30 (ECFP), 510/20 (EGFP) and 550/30 (EYFP) bandpasses; 458 nm laser blocking filters. Results: All three fluorescent proteins when expressed individually or in combination in living cells were excited by the 458 nm laser line and theircorresponding signals could be electronically compensated in real time. Conclusions: This method demonstrates the detection of three fluorescent proteins expressed simultaneously in living cells using single laser excitation and is applicable for use on flow cytometers equipped with a tunable argon ion laser. (C) 1999 Wiley-Liss,Inc.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 04/12/20 alle ore 09:54:11