Time-gated FRET nanoassemblies for rapid and sensitive intra- and extracellular fluorescence imaging

Time-gated Förster resonance energy transfer (FRET) using the unique material combination of long-lifetime terbium complexes (Tb) and semiconductor quantum dots (QDs) provides many advantages for highly sensitive and multiplexed biosensing. Although time-gated detection can efficiently suppress samp...

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Bibliographic Details
Published in:Science advances 2016-06, Vol.2 (6), p.e1600265
Main Authors: Afsari, Hamid Samareh, Cardoso Dos Santos, Marcelina, Lindén, Stina, Chen, Ting, Qiu, Xue, van Bergen En Henegouwen, Paul M P, Jennings, Travis L, Susumu, Kimihiro, Medintz, Igor L, Hildebrandt, Niko, Miller, Lawrence W
Format: Article
Language:English
Subjects:
Biosensors
Cell Line
Cell-Penetrating Peptides
Chemical Sciences
Endocytosis
Extracellular Space
Fluorescence Resonance Energy Transfer - methods
Humans
Intracellular Space
Life Sciences
Microinjections
Molecular Imaging - methods
Molecular Imaging - standards
Nanostructures - chemistry
Optical Imaging - methods
Optical Imaging - standards
Physics
Quantum Dots
SciAdv r-articles
Semiconductors
Sensitivity and Specificity
Single-Domain Antibodies
Terbium
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Summary:Time-gated Förster resonance energy transfer (FRET) using the unique material combination of long-lifetime terbium complexes (Tb) and semiconductor quantum dots (QDs) provides many advantages for highly sensitive and multiplexed biosensing. Although time-gated detection can efficiently suppress sample autofluorescence and background fluorescence from directly excited FRET acceptors, Tb-to-QD FRET has rarely been exploited for biomolecular imaging. We demonstrate Tb-to-QD time-gated FRET nanoassemblies that can be applied for intra- and extracellular imaging. Immunostaining of different epitopes of the epidermal growth factor receptor (EGFR) with Tb- and QD-conjugated antibodies and nanobodies allowed for efficient Tb-to-QD FRET on A431 cell membranes. The broad usability of Tb-to-QD FRET was further demonstrated by intracellular Tb-to-QD FRET and Tb-to-QD-to-dye FRET using microinjection as well as cell-penetrating peptide-mediated endocytosis with HeLa cells. Effective brightness enhancement by FRET from several Tb to the same QD, the use of low nanomolar concentrations, and the quick and sensitive detection void of FRET acceptor background fluorescence are important advantages for advanced intra- and extracellular imaging of biomolecular interactions.
ISSN:2375-2548
2375-2548
DOI:10.1126/sciadv.1600265