Time-gated FRET nanoassemblies for rapid and sensitive intra- and extracellular fluorescence imaging
journal contributionposted on 17.12.2021, 16:39 by HS Afsari, MC Dos Santos, S Lindén, T Chen, X Qiu, PMP Van Bergen en Henegouwen, TL Jennings, K Susumu, IL Medintz, N Hildebrandt, Lawrence MillerLawrence Miller
Time-gated Förster resonance energy transfer (FRET) using the uniquematerial 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 fromseveral Tb to the sameQD, the use of lownanomolar 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.
Targeted Lanthanide Contrast Agents for in Cellulo Single Molecule Imaging | Funder: National Institutes of Health (National Institute of General Medical Sciences) | Grant ID: R01GM081030
CitationAfsari, H. S., Dos Santos, M. C., Lindén, S., Chen, T., Qiu, X., Van Bergen en Henegouwen, P. M. P., Jennings, T. L., Susumu, K., Medintz, I. L., Hildebrandt, N.Miller, L. W. (2016). Time-gated FRET nanoassemblies for rapid and sensitive intra- and extracellular fluorescence imaging. Science Advances, 2(6), e1600265-. https://doi.org/10.1126/sciadv.1600265
PublisherAmerican Association for the Advancement of Science (AAAS)
Read the peer-reviewed publication
NanotechnologyBioengineering1.1 Normal biological development and functioningFRET relayscell penetrating peptidescytosolendocytosisimmunostainingmicroinjectionnanobodiesquantum dotsterbiumCell LineCell-Penetrating PeptidesEndocytosisExtracellular SpaceFluorescence Resonance Energy TransferHumansIntracellular SpaceMicroinjectionsMolecular ImagingNanostructuresOptical ImagingQuantum DotsSemiconductorsSensitivity and SpecificitySingle-Domain AntibodiesTerbium