scholarly journals Quantum dot conjugated nanobodies for multiplex imaging of protein dynamics at synapses

Nanoscale ◽  
2018 ◽  
Vol 10 (21) ◽  
pp. 10241-10249 ◽  
Author(s):  
Souvik Modi ◽  
Nathalie F. Higgs ◽  
David Sheehan ◽  
Lewis D. Griffin ◽  
Josef T. Kittler
Keyword(s):  
Cell Membrane ◽  
Protein Dynamics ◽  
Particle Tracking ◽  
Single Particle ◽  
Ex Vivo ◽  
Optical Probe ◽  
Single Particle Tracking ◽  
Specific Probe ◽  
Diffusion Dynamics

An anti-GFP nanobody conjugated QD optical probe was applied to study single particle trackingin vitroandex vivo. This small, highly specific probe recognized GFP when expressed at the cell membrane and reported diffusion dynamics of the underlying target protein.

scholarly journals Functional Characterization of an Electromagnetic Perceptive Protein

2020 ◽  
Author(s):  
Sunayana Mitra ◽  
Carlo Barnaba ◽  
Jens Schmidt ◽  
Galit Pelled ◽  
Assaf A. Gilad
Keyword(s):  
Particle Tracking ◽  
Single Particle ◽  
Total Internal Reflection ◽  
Magnetic Stimulation ◽  
New Technologies ◽  
Cellular Function ◽  
Single Particle Tracking ◽  
Internal Reflection ◽  
Total Internal Reflection Fluorescence ◽  
Diffusion Dynamics

AbstractMagnetoreception, the response to geomagnetic fields is a well described phenomenon in nature. However, it is likely that convergent evolution led to different mechanisms in different organisms. One intriguing example is the unique Electromagnetic Perceptive Gene (EPG) from the glass catfish Kryptopterus vitreolus, that can remotely control cellular function, upon magnetic stimulation in in-vitro and in animal models. Here, we report for the first time the cellular location and orientation of the EPG protein. We utilized a differential labelling technique in determining that the EPG protein is a membrane anchored protein with an N-terminal extracellular domain. The kinetics and diffusion dynamics of the EPG protein in response to magnetic stimulation was also elucidated using single particle imaging and tracking. Pulse chase labelling and Total Internal Reflection Fluorescence (TIRF) imaging revealed an increase in EPG kinetics post magnetic stimulation activation at a single particle level. Trajectory analysis show notably different EPG protein kinetics before and after magnetic stimulation in both 2 (free vs bound particle) and 3 state (free vs intermediate vs bound particle) tracking models. These data serve to provide additional information that support and understand the underlying biophysical mechanisms behind EPG activation by magnetic stimulation. In conclusion, our results provide evidence for the basis of magnetoreception in EPG protein that would aid in future studies that seek to understand this novel mechanism. This study is important for understanding the phenomenon of magnetoreception as well as developing new technologies for magnetogenetics – the utilization of electromagnetic fields to remotely control cellular function.Graphical TOCElucidation of magnetoreception in a fish derived Electromagnetic Perceptive Gene (EPG), using genetic tagging and single particle tracking with Total Internal Reflection Fluorescence (TIRF) suggests changes in kinetics of membranal motion upon stimulation by magnetic field.

Correlation between the translational and rotational diffusion of rod-shaped nanocargo on a lipid membrane revealed by single-particle tracking

Nanoscale ◽  
2019 ◽  
Vol 11 (20) ◽  
pp. 10080-10087 ◽  
Author(s):  
Liangna He ◽  
Yiliang Li ◽  
Lin Wei ◽  
Zhongju Ye ◽  
Hua Liu ◽  
...  
Keyword(s):  
Particle Tracking ◽  
Single Particle ◽  
Theoretical Basis ◽  
Rational Design ◽  
Lipid Membrane ◽  
Rotational Diffusion ◽  
Single Particle Tracking ◽  
Cellular Translocation ◽  
Diffusion Dynamics ◽  
Translational And Rotational Diffusion

Revealing the diffusion dynamics of nanoparticles on a lipid membrane plays an important role in a better understanding of the cellular translocation process and provides a theoretical basis for the rational design of delivery cargo.

scholarly journals Cega: A Single Particle Segmentation Algorithm to Identify Moving Particles in a Noisy System

2020 ◽  
Author(s):  
Erin M. Masucci ◽  
Peter K. Relich ◽  
E. Michael Ostap ◽  
Erika L. F. Holzbaur ◽  
Melike Lakadamyali
Keyword(s):  
Particle Tracking ◽  
Single Particle ◽  
Molecular Motors ◽  
Single Particle Tracking ◽  
Particle Detection ◽  
Identification Rate ◽  
Detection Techniques ◽  
Noticeable Improvement

ABSTRACTImprovements to particle tracking algorithms are required to effectively analyze the motility of biological molecules in complex or noisy systems. A typical single particle tracking (SPT) algorithm detects particle coordinates for trajectory assembly. However, particle detection filters fail for datasets with low signal-to-noise levels. When tracking molecular motors in complex systems, standard techniques often fail to separate the fluorescent signatures of moving particles from background noise. We developed an approach to analyze the motility of kinesin motor proteins moving along the microtubule cytoskeleton of extracted neurons using the Kullback-Leibler (KL) divergence to identify regions where there are significant differences between models of moving particles and background signal. We tested our software on both simulated and experimental data and found a noticeable improvement in SPT capability and a higher identification rate of motors as compared to current methods. This algorithm, called Cega, for ‘find the object’, produces data amenable to conventional blob detection techniques that can then be used to obtain coordinates for downstream SPT processing. We anticipate that this algorithm will be useful for those interested in tracking moving particles in complex in vitro or in vivo environments.

scholarly journals Hop diffusion in the cell membrane is universally found as revealed by single fluorophore imaging and single particle tracking

2003 ◽  
Vol 43 (supplement) ◽  
pp. S157
Author(s):  
K. Iwasawa ◽  
T. Fujiwara ◽  
J. Kondo ◽  
K. Murase ◽  
H. Yamashita ◽  
...  
Keyword(s):  
Cell Membrane ◽  
Particle Tracking ◽  
Single Particle ◽  
Single Particle Tracking

scholarly journals Single-Particle Tracking for the Quantification of Membrane Protein Dynamics in Living Plant Cells

2018 ◽  
Vol 11 (11) ◽  
pp. 1315-1327 ◽  
Author(s):  
Yaning Cui ◽  
Meng Yu ◽  
Xiaomin Yao ◽  
Jingjing Xing ◽  
Jinxing Lin ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Protein Dynamics ◽  
Particle Tracking ◽  
Single Particle ◽  
Plant Cells ◽  
Single Particle Tracking ◽  
Living Plant ◽  
Membrane Protein Dynamics

scholarly journals Spot-On: robust model-based analysis of single-particle tracking experiments

2017 ◽  
Author(s):  
Anders S Hansen ◽  
Maxime Woringer ◽  
Jonathan B Grimm ◽  
Luke D Lavis ◽  
Robert Tjian ◽  
...  
Keyword(s):  
Single Molecule ◽  
Particle Tracking ◽  
Single Particle ◽  
Motion Blur ◽  
Single Particle Tracking ◽  
Modeling Framework ◽  
Diffusion Constants ◽  
Diffusion Dynamics ◽  
Wide Range ◽  
And Diffusion

ABSTRACTSingle-particle tracking (SPT) has become an important method to bridge biochemistry and cell biology since it allows direct observation of protein binding and diffusion dynamics in live cells. However, accurately inferring information from SPT studies is challenging due to biases in both data analysis and experimental design. To address analysis bias, we introduce “Spot-On”, an intuitive web-interface. Spot-On implements a kinetic modeling framework that accounts for known biases, including molecules moving out-of-focus, and robustly infers diffusion constants and subpopulations from pooled single-molecule trajectories. To minimize inherent experimental biases, we implement and validate stroboscopic photo-activation SPT (spaSPT), which minimizes motion-blur bias and tracking errors. We validate Spot-On using experimentally realistic simulations and show that Spot-On outperforms other methods. We then apply Spot-On to spaSPT data from live mammalian cells spanning a wide range of nuclear dynamics and demonstrate that Spot-On consistently and robustly infers subpopulation fractions and diffusion constants.IMPACT STATEMENTSpot-On is an easy-to-use website that makes a rigorous and bias-corrected modeling framework for analysis of single-molecule tracking experiments available to all.

scholarly journals Analysis of α3 GlyR single particle tracking in the cell membrane

2014 ◽  
Vol 1843 (3) ◽  
pp. 544-553 ◽  
Author(s):  
Kristof Notelaers ◽  
Susana Rocha ◽  
Rik Paesen ◽  
Nick Smisdom ◽  
Ben De Clercq ◽  
...  
Keyword(s):  
Cell Membrane ◽  
Particle Tracking ◽  
Single Particle ◽  
Single Particle Tracking
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