Tracking Individual Proteins in Living Cells Using Single Quantum Dot Imaging

Courty, Sébastien; Bouzigues, Cédric; Luccardini, Camilla; Ehrensperger, Marie-Virginie; Bonneau, Stéphane; Dahan, Maxime

doi:10.1016/s0076-6879(06)14012-4

Cited by 33 publications

(25 citation statements)

References 30 publications

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“…QDs were selected for tracking analysis only if they were localized to photoreceptor terminals, exhibited a small size (≤4 pixels) and showed intermittent blinking; these criteria are consistent with labeling by a single quantum dot (Alcor et al, 2009). The location of a QD can be determined with precision exceeding the diffraction limit by fitting the fluorescence profile with a Gaussian point spread function (Courty et al, 2006). To estimate pointing accuracy, we measured the standard deviation of displacements exhibited by QDs immobilized in vacuum grease and found that they averaged 96 nm on the upright microscope (11 QDs, 100 measurements/QD) and 15 nm on the inverted microscope (10 QDs).…”

Section: Methodsmentioning

confidence: 99%

Lateral Mobility of Presynaptic L-Type Calcium Channels at Photoreceptor Ribbon Synapses

2011

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At most synapses, presynaptic Ca2+ channels are positioned near vesicle release sites, and increasing this distance reduces synaptic strength. We examined the lateral membrane mobility of presynaptic L-type Ca2+ channels at photoreceptor ribbon synapses of the tiger salamander (Ambystoma tigrinum) retina. Movements of individual Ca2+ channels were tracked by coupling quantum dots to an antibody against the extracellular α2δ4 Ca2+ channel subunit. α2δ4 antibodies labeled photoreceptor terminals and co-localized with antibodies to synaptic vesicle protein SV2 and Ca2+ channel CaV1.4 α1 subunits. The results show that Ca2+ channels are dynamic and move within a confined region beneath the synaptic ribbon. The size of this confinement area is regulated by actin and membrane cholesterol. Fusion of nearby synaptic vesicles caused jumps in Ca2+ channel position, propelling them towards the outer edge of the confinement domain. Channels rebounded rapidly towards the center. Thus, although CaV channels are mobile, molecular scaffolds confine them beneath the ribbon to maintain neurotransmission even at high release rates.

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Section: Methodsmentioning

confidence: 99%

Lateral Mobility of Presynaptic L-Type Calcium Channels at Photoreceptor Ribbon Synapses

2011

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“…These applications have inspired the use QDs for monitoring other plasma membrane proteins such as integrins [50,66], tyrosine kinases [67,68], G-protein coupled receptors [69], and membrane lipids associated with apoptosis [70,71]. As well, detailed procedures for receptor labeling and visualization of receptor dynamics with QDs have recently been published [72,73], and new techniques to label plasma membrane proteins using versatile molecular biology methods have been developed [74,75].…”

Section: Imaging and Tracking Of Membrane Receptorsmentioning

confidence: 99%

Bioconjugated quantum dots for in vivo molecular and cellular imaging☆

Smith

Duan

Mohs

et al. 2008

Advanced Drug Delivery Reviews

1,086

760

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Semiconductor quantum dots (QDs) are tiny light-emitting particles on the nanometer scale, and are emerging as a new class of fluorescent labels for biology and medicine. In comparison with organic dyes and fluorescent proteins, they have unique optical and electronic properties, with size-tunable light emission, superior signal brightness, resistance to photobleaching, and broad absorption spectra for simultaneous excitation of multiple fluorescence colors. QDs also provide a versatile nanoscale scaffold for designing multifunctional nanoparticles with both imaging and therapeutic functions. When linked with targeting ligands such as antibodies, peptides or small molecules, QDs can be used to target tumor biomarkers as well as tumor vasculatures with high affinity and specificity. Here we discuss the synthesis and development of state-of-the-art QD probes and their use for molecular and cellular imaging. We also examine key issues for in vivo imaging and therapy, such as nanoparticle biodistribution, pharmacokinetics, and toxicology.

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“…These advantageous properties overcome difficulties in photobleaching that limit fluorophore imaging time allowing for acquisition of biological events over long timescales and contribute to the QD’s utility as an ultrasensitive detection probe for SPT. Moreover, QD-SPT generates quantifiable dynamic information regarding diffusional properties, co-localization, and spatial and temporal heterogeneity of molecules inside living cells that conventional fluorescence and biochemical methods cannot capture (Courty et al 2006a; Cognet et al 2014; Breger et al 2014). …”

Section: Qd-enabled Studies Of Single Molecule Behavior In Living Cellsmentioning

confidence: 99%

“…Following labeling and validation of the QD probe, a sequence of fluorescence images are acquired in time lapse to capture the biological event. The temporal resolution achieved through this acquisition will usually be limited by the camera readout time rather than by the brightness of the probe (Courty et al 2006a). Finally, biological information is extracted from the recorded trajectories through single particle tracking to yield measurements such as diffusion coefficient and velocity that provide dynamic information about the molecule of interest (Bannai et al 2006).…”

Section: Qd-enabled Studies Of Single Molecule Behavior In Living Cellsmentioning

confidence: 99%

Quantum dots for quantitative imaging: from single molecules to tissue

Lam

Hatch

et al. 2015

Cell Tissue Res

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Since their introduction to biological imaging, quantum dots (QDs) have progressed from a little known, but attractive technology to one that has gained broad application in many areas of biology. The versatile properties of these fluorescent nanoparticles have allowed investigators to conduct biological studies with extended spatiotemporal capabilities that were previously not possible. In this review, we focus on QD applications that provide enhanced quantitative information on protein dynamics and localization, including single particle tracking (SPT) and immunohistochemistry (IHC), and finish by examining prospects of upcoming applications, such as correlative light and electron microscopy (CLEM) and super-resolution. Advances in single molecule imaging, including multi-color and 3D QD tracking, have provided new insights into the mechanisms of cell signaling and protein trafficking. New forms of QD tracking in vivo have allowed for observation of biological processes with molecular level resolution in the physiological context of the whole animal. Further methodological development of multiplexed QD-based immunohistochemistry assays are allowing more quantitative analysis of key proteins in tissue samples. These advances highlight the unique quantitative data sets that QDs can provide to further our understanding of biological and disease processes.

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Tracking Individual Proteins in Living Cells Using Single Quantum Dot Imaging

Cited by 33 publications

References 30 publications

Lateral Mobility of Presynaptic L-Type Calcium Channels at Photoreceptor Ribbon Synapses

Lateral Mobility of Presynaptic L-Type Calcium Channels at Photoreceptor Ribbon Synapses

Bioconjugated quantum dots for in vivo molecular and cellular imaging☆

Quantum dots for quantitative imaging: from single molecules to tissue

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