scholarly journals Field-Controlled Charge Separation in a Conductive Matrix at the Single-Molecule Level: Toward Controlling Single-Molecule Fluorescence Intermittency

ACS Omega ◽  
2016 ◽  
Vol 1 (6) ◽  
pp. 1383-1392 ◽  
Author(s):  
Koen Kennes ◽  
Peter Dedecker ◽  
James A. Hutchison ◽  
Eduard Fron ◽  
Hiroshi Uji-i ◽  
...  
Keyword(s):  
Single Molecule ◽  
Charge Separation ◽  
Single Molecule Fluorescence ◽  
Single Molecule Level ◽  
Fluorescence Intermittency

Styryl dye formation promoted by catalytic centers of piperazine bound to a silica surface traced by single molecule fluorescence microscopy

2017 ◽  
Vol 19 (31) ◽  
pp. 20984-20990 ◽  
Author(s):  
Aline M. Lino ◽  
Marcelo H. Gehlen
Keyword(s):  
Fluorescence Microscopy ◽  
Single Molecule ◽  
Silica Surface ◽  
Knoevenagel Condensation ◽  
Fluorescent Dye ◽  
New Method ◽  
Single Molecule Fluorescence ◽  
Single Molecule Level ◽  
Styryl Dye ◽  
Single Molecule Fluorescence Microscopy

Knoevenagel condensation producing a fluorescent dye is studied at a single molecule level establishing a new method to investigate the molecular rate of amino catalysis.

scholarly journals Stoichiometric Analysis of Protein Complexes by Cell Fusion and Single Molecule Imaging

2020 ◽  
Author(s):  
Avtar Singh ◽  
Alexander L. Van Slyke ◽  
Maria Sirenko ◽  
Alexander Song ◽  
Paul J. Kammermeier ◽  
...  
Keyword(s):  
Single Molecule ◽  
Fluorescence Correlation Spectroscopy ◽  
Protein Complexes ◽  
Correlation Spectroscopy ◽  
Single Molecule Fluorescence ◽  
Single Molecule Level ◽  
Fluorescence Correlation ◽  
Cellular Environment ◽  
Low Expression

ABSTRACTThe composition, stoichiometry and interactions of supramolecular protein complexes are a critical determinant of biological function. Several techniques have been developed to study molecular interactions and quantify subunit stoichiometry at the single molecule level; however, these typically require artificially low expression levels to achieve the low fluorophore concentration required for single molecule imaging, or use of detergent isolation of complexes that may perturb native subunit interactions. Here we present an alternative approach where protein complexes are assembled at physiological concentrations and subsequently diluted in situ for single-molecule level observations while preserving them in a near-native cellular environment. We show that coupling this in situ dilution strategy with single molecule techniques such as in vivo Fluorescence Correlation Spectroscopy (FCS), bleach step counting for quantifying protein complex stoichiometry, and two-color single molecule colocalization, improves the quality of data obtained using these single molecule fluorescence methods. Single Protein Recovery After Dilution (SPReAD) is a simple and versatile means of extending the concentration range of single molecule measurements into the cellular regime while minimizing potential artifacts and perturbations of protein complex stoichiometry.SIGNIFICANCE STATEMENTQuantifying the composition and stoichiometry of protein complexes in live cells is critical to understanding mechanisms involved in their function. Here we detail a method in which protein complexes are assembled intracellularly at physiological concentrations, but then diluted to levels suitable for single-molecule fluorescence observations while still within a cellular environment. The technique permits the use of common single molecule analysis techniques such as stepwise photobleaching quantification and fluorescence correlation spectroscopy to determine stoichiometry and functional interactions while avoiding artifacts that may occur from the use of detergent isolation methods or from the artificially low expression levels sometimes used to attain single molecule observation levels.

Single-molecule fluorescence to study molecular motors

2007 ◽  
Vol 40 (1) ◽  
pp. 87-111 ◽  
Author(s):  
Hyokeun Park ◽  
Erdal Toprak ◽  
Paul R. Selvin
Keyword(s):  
Fluorescence Imaging ◽  
Single Molecule ◽  
Molecular Motors ◽  
Atp Hydrolysis ◽  
Imaging Techniques ◽  
Single Molecule Fluorescence ◽  
New Techniques ◽  
Single Molecule Level ◽  
Living Organisms ◽  
Run Lengths

AbstractMolecular motors, which use energy from ATP hydrolysis to take nanometer-scale steps with run-lengths on the order of micrometers, have important roles in areas such as transport and mitosis in living organisms. New techniques have recently been developed to measure these small movements at the single-molecule level. In particular, fluorescence imaging has contributed to the accurate measurement of this tiny movement. We introduce three single-molecule fluorescence imaging techniques which can find the position of a fluorophore with accuracy in the range of a few nanometers. These techniques are named after Hollywood animation characters: Fluorescence Imaging with One Nanometer Accuracy (FIONA), Single-molecule High-REsolution Colocalization (SHREC), and Defocused Orientation and Position Imaging (DOPI). We explain new understanding of molecular motors obtained from measurements using these techniques.

scholarly journals Exponential and Power-Law Kinetics in Single-Molecule Fluorescence Intermittency†

2004 ◽  
Vol 108 (29) ◽  
pp. 10445-10450 ◽  
Author(s):  
Matthias Haase ◽  
Christian G. Hübner ◽  
E. Reuther ◽  
A. Herrmann ◽  
K. Müllen ◽  
...  
Keyword(s):  
Single Molecule ◽  
Power Law ◽  
Single Molecule Fluorescence ◽  
Power Law Kinetics ◽  
Fluorescence Intermittency
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