Fluorescence single-molecule counting assays for protein quantification using epi-fluorescence microscopy with quantum dots labeling

2010 ◽  
Vol 662 (2) ◽  
pp. 170-176 ◽  
Author(s):  
Dafeng Jiang ◽  
Chunxia Liu ◽  
Lei Wang ◽  
Wei Jiang
Keyword(s):  
Quantum Dots ◽  
Fluorescence Microscopy ◽  
Single Molecule ◽  
Protein Quantification

A Single-Molecule Homogeneous Immunoassay by Counting Spatially “Overlapping” Two-Color Quantum Dots with Wide-Field Fluorescence Microscopy

ACS Sensors ◽  
2018 ◽  
Vol 3 (12) ◽  
pp. 2644-2650 ◽  
Author(s):  
Xiaojun Liu ◽  
Conghui Huang ◽  
Chenghua Zong ◽  
Aiye Liang ◽  
Zhangjian Wu ◽  
...  
Keyword(s):  
Quantum Dots ◽  
Fluorescence Microscopy ◽  
Single Molecule ◽  
Wide Field ◽  
Homogeneous Immunoassay

scholarly journals Distance dependence of near-field fluorescence enhancement and quenching of single quantum dots

2011 ◽  
Vol 2 ◽  
pp. 645-652 ◽  
Author(s):  
Volker Walhorn ◽  
Jan Paskarbeit ◽  
Heinrich Gotthard Frey ◽  
Alexander Harder ◽  
Dario Anselmetti
Keyword(s):  
Quantum Dots ◽  
Fluorescence Microscopy ◽  
Single Molecule ◽  
Dipolar Coupling ◽  
Near Field ◽  
Fluorescence Emission ◽  
Field Enhancement ◽  
Single Quantum ◽  
Distance Dependence ◽  
Single Quantum Dots

In fluorescence microscopy and spectroscopy, energy transfer processes between single fluorophores and fluorophore quencher pairs play an important role in the investigation of molecular distances or orientations. At distances larger than about 3 nm these effects originate predominantly from dipolar coupling. As these experiments are commonly performed in homogenous media, effects at the interface boundaries can be neglected. Nevertheless, the combination of such assays with single-molecule manipulation techniques such as atomic force microscopy (AFM) requires a detailed understanding of the influence of interfaces on dipolar coupling effects. In the presented work we used a combined total internal reflection fluorescence microscopy (TIRFM)–AFM setup to elucidate this issue. We measured the fluorescence emission emanating from single quantum dots as a function of distance from the apex of a gold-coated cantilever tip. As well as fluorescence quenching at close proximity to the tip, we found a nonlinear and nonmonotonic distance dependence of the fluorescence emission. To confirm and interpret our findings we performed calculations on the basis of a simplified multiple multipole (MMP) approach, which successfully supports our experimental data. Moreover, we revealed and quantified the influence of interfering processes such as field enhancement confined at interface boundaries, mirror dipoles and (resonant) dipolar coupling.

scholarly journals Low-Light Photodetectors for Fluorescence Microscopy

2021 ◽  
Vol 11 (6) ◽  
pp. 2773
Author(s):  
Hiroaki Yokota ◽  
Atsuhito Fukasawa ◽  
Minako Hirano ◽  
Toru Ide
Keyword(s):  
Fluorescence Microscopy ◽  
Fluorescence Imaging ◽  
Single Molecule ◽  
Science Studies ◽  
Single Photon ◽  
Laser Scanning Microscopy ◽  
Oxide Semiconductor ◽  
Wide Field ◽  
Single Molecule Fluorescence ◽  
Low Light

Over the years, fluorescence microscopy has evolved and has become a necessary element of life science studies. Microscopy has elucidated biological processes in live cells and organisms, and also enabled tracking of biomolecules in real time. Development of highly sensitive photodetectors and light sources, in addition to the evolution of various illumination methods and fluorophores, has helped microscopy acquire single-molecule fluorescence sensitivity, enabling single-molecule fluorescence imaging and detection. Low-light photodetectors used in microscopy are classified into two categories: point photodetectors and wide-field photodetectors. Although point photodetectors, notably photomultiplier tubes (PMTs), have been commonly used in laser scanning microscopy (LSM) with a confocal illumination setup, wide-field photodetectors, such as electron-multiplying charge-coupled devices (EMCCDs) and scientific complementary metal-oxide-semiconductor (sCMOS) cameras have been used in fluorescence imaging. This review focuses on the former low-light point photodetectors and presents their fluorescence microscopy applications and recent progress. These photodetectors include conventional PMTs, single photon avalanche diodes (SPADs), hybrid photodetectors (HPDs), in addition to newly emerging photodetectors, such as silicon photomultipliers (SiPMs) (also known as multi-pixel photon counters (MPPCs)) and superconducting nanowire single photon detectors (SSPDs). In particular, this review shows distinctive features of HPD and application of HPD to wide-field single-molecule fluorescence detection.

scholarly journals Structural Contributions to Hydrodynamic Diameter for Quantum Dots Optimized for Live-Cell Single-Molecule Tracking

2018 ◽  
Vol 122 (30) ◽  
pp. 17406-17412 ◽  
Author(s):  
Janet Y. Sheung ◽  
Pinghua Ge ◽  
Sung Jun Lim ◽  
Sang Hak Lee ◽  
Andrew M. Smith ◽  
...  
Keyword(s):  
Quantum Dots ◽  
Single Molecule ◽  
Live Cell ◽  
Hydrodynamic Diameter ◽  
Single Molecule Tracking

scholarly journals Total Internal Reflection Fluorescence Microscopy (TIRFM) – novel techniques and applications

2020 ◽  
Vol 8 (11) ◽  
Author(s):  
Verena Richter ◽  
Michael Wagner ◽  
Herbert Schneckenburger
Keyword(s):  
Fluorescence Microscopy ◽  
Single Molecule ◽  
Total Internal Reflection ◽  
Plasma Membranes ◽  
Focal Adhesions ◽  
Internal Reflection ◽  
Total Internal Reflection Fluorescence ◽  
Thin Layers ◽  
Structured Illumination ◽  
Structured Illumination Microscopy

Total Internal Reflection Fluorescence Microscopy (TIRFM) has been established almost 40 years ago for studies of plasma membranes or membrane proximal sites of living cells. The method is based on light incidence at an angle above the critical angle of total internal reflection and generation of an evanescent electromagnetic field penetrating about 100 nm into a sample and permitting selective excitation of membrane proximal fluorophores. Two techniques are presented here: prism-type TIRFM and objective-type TIRFM with high aperture microscope objective lenses. Furthermore, numerous applications are summarized, e.g. measurement of focal adhesions, cell-substrate topology, endocytosis or exocytosis of vesicles as well as single molecule detection within thin layers. Finally, highly innovative combinations of TIRFM with Förster Resonance Energy Transfer (FRET) measurements as well as with Structured Illumination Microscopy (SIM) and fluorescence reader technologies are presented.

Fabrication of Low Cost 1D CdSe Nanowires using Near-field Electrospinning

2011 ◽  
Vol 1302 ◽  
Author(s):  
Leroy Magwood ◽  
Binil Starly
Keyword(s):  
Quantum Dots ◽  
Fluorescence Microscopy ◽  
Fiber Diameter ◽  
Low Cost ◽  
Near Field ◽  
Light Attenuation ◽  
Dimethyl Formamide ◽  
Cdse Quantum Dot ◽  
The Matrix ◽  
Cdse Nanowires

ABSTRACTWell-aligned, 1D CdSe quantum dot (QD) fibers (0.3μm to 2.5μm) containing up to 20wt% fluorescent quantum dots (QDs) were prepared by near-field electrospinning (NFES) process. Electrospun solutions were prepared using PVAc as the matrix polymer, dimethyl formamide (DMF) solvent and colloidal QDs in chloroform (CHCl3). The diameter of the fibers decreased as the ratio of DMF/CHCl3 is varied. QDs showed good dispersion and a linear relationship between QD loading and fiber diameter, as determined by the morphology measurements taken using TEM and SEM, respectively. Fluorescence microscopy shows that there is light attenuation throughout the fibers. Results also show that the NFES process may be used as a method to create aligned, 1D fibers of QDs and potentially other nanofibers.

scholarly journals HDL particles incorporate into lipid bilayers – a combined AFM and single molecule fluorescence microscopy study

2017 ◽  
Vol 7 (1) ◽  
Author(s):  
Birgit Plochberger ◽  
Clemens Röhrl ◽  
Johannes Preiner ◽  
Christian Rankl ◽  
Mario Brameshuber ◽  
...  
Keyword(s):  
Fluorescence Microscopy ◽  
Lipid Bilayers ◽  
Single Molecule ◽  
Microscopy Study ◽  
Single Molecule Fluorescence ◽  
Single Molecule Fluorescence Microscopy

scholarly journals Quantification of Single-Molecule FRET between Quantum Dots and Organic Dyes

2018 ◽  
Vol 114 (3) ◽  
pp. 684a
Author(s):  
Nooshin Shatery Nejad ◽  
Candice M. Etson
Keyword(s):  
Quantum Dots ◽  
Single Molecule ◽  
Organic Dyes ◽  
Single Molecule Fret
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