Re-evaluation of biotin-streptavidin conjugation in Förster resonance energy transfer applications.
J Biomed Opt
; 19(8): 085008, 2014 Aug.
Article
em En
| MEDLINE
| ID: mdl-25162908
ABSTRACT
Bioaffinity conjugation between streptavidin (SA) and biotin has been widely used to link donors and acceptors for investigating the distance-dependent Förster resonance energy transfer (FRET). When studying a commonly used FRET system of (QD-SA)-(biotin-DNA-dye) [donor quantum dot (QD); acceptor small organic fluorescent dye; and linker deoxyribose nucleic acid (DNA) molecule via SA-biotin conjugation], however, a contradictory finding was recently reported in the literature. It was found that the FRET lost its dependence on the number of DNA base pairs when using a phosphate-buffered saline (PBS) solution. We found that the conflicted results were caused by the ionic strength of the adopted buffer solutions. Our results suggest that the dependent FRET on the number of DNA bases is favorable in a low-ionic-strength buffer, whereas in relatively high-ionic-strength buffers, the FRET loses the DNA length dependence. We propose that the independence is mainly caused by the conformational change of DNA molecules from a stretched to a coiled mode when the cations in the high-ionic-strength buffer neutralize the negatively charged backbone of DNA molecules, thereby bringing the acceptors close to the donors.
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Biotina
/
DNA
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Imunoensaio
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Artefatos
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Estreptavidina
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Transferência Ressonante de Energia de Fluorescência
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Pontos Quânticos
Tipo de estudo:
Diagnostic_studies
Idioma:
En
Ano de publicação:
2014
Tipo de documento:
Article