A Two-Step Method for smFRET Data Analysis

Chen, Jixin; Pyle, Joseph R.; Piecco, Kurt Waldo Sy; Kolomeisky, Anatoly B.; Landes, Christy F.

doi:10.1021/acs.jpcb.6b05697

“…The probability of n ph incident photons to be detected on the detector pixel ( x , y ) during a frame l is given by a Poisson distribution P ( n ph ; μ ph = I ( x , y , l ) D/A characterized by the mean number of incident photons μ ph . The simulation of sole photon noise is commonly used for the simulation of single photon detection applications [ 19 , 45 , 59 – 62 ]. (ii) The second noise contribution is the camera shot noise σ pe common to all (EM)CCDs or sCMOS [ 42 , 43 ].…”

Section: Simulation Of Smvsmentioning

confidence: 99%

“…Instead, individual researchers face a host of different data analysis strategies developed by different groups [ 12 , 14 – 17 ]. Therein, SMV simulations in camera-based SM detection have been reported in the context of kinetic/dwell-time analysis, where method-specific kinetic rates are determined and evaluated against their simulated ground truth, usually as a function of the SNR [ 12 , 14 , 18 , 19 ]. Although these studies describe the simulation process used, recovering video simulation parameter (VSP) values from the manuscript or supplementary material to reproduce the simulated SMV is typically not straightforward.…”

Section: Introductionmentioning

confidence: 99%

Simulations of camera-based single-molecule fluorescence experiments

Börner

¹

,

Kowerko

²

,

Hadzic

³

et al. 2018

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Single-molecule microscopy has become a widely used technique in (bio)physics and (bio)chemistry. A popular implementation is single-molecule Förster Resonance Energy Transfer (smFRET), for which total internal reflection fluorescence microscopy is frequently combined with camera-based detection of surface-immobilized molecules. Camera-based smFRET experiments generate large and complex datasets and several methods for video processing and analysis have been reported. As these algorithms often address similar aspects in video analysis, there is a growing need for standardized comparison. Here, we present a Matlab-based software (MASH-FRET) that allows for the simulation of camera-based smFRET videos, yielding standardized data sets suitable for benchmarking video processing algorithms. The software permits to vary parameters that are relevant in cameras-based smFRET, such as video quality, and the properties of the system under study. Experimental noise is modeled taking into account photon statistics and camera noise. Finally, we survey how video test sets should be designed to evaluate currently available data analysis strategies in camera-based sm fluorescence experiments. We complement our study by pre-optimizing and evaluating spot detection algorithms using our simulated video test sets.

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“…A single CPU is used for all simulations. A previously coded exponential regression fitting algorithm jcfit 45 (Github) is used to fit the curves. The two major random functions to generate the step motion of each molecule are from MATLAB, rand() creating evenly distributed random numbers, and randn() generating Gaussian distributed random numbers.…”

Section: Methodsmentioning

confidence: 99%

Stochastic Adsorption of Diluted Solute Molecules at Interfaces

Chen

¹

2021

Preprint

Self Cite

1

0

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<div> <p>Here an analytical solution of Fick’s 2<sup>nd</sup> law is used to predict the diffusion and the stochastic adsorption of single diluted solute molecules on flat and patterned surfaces. The equations are then compared to the results of several numerical Monte Carlo simulations using a random walk model. The 1D diffusion simulations clarify that the dependence of the solute-surface collision rate on the observation-time (measurement time resolution) is because of the multiple collisions of the same molecules over different time regions. It also surprisingly suggests that due to the self-mimetic fractal function of diffusion, the equation should be corrected by a factor of two. The absorption rate of solute on an adsorptive surface is found to follow a power-law decay function due to an evolving concentration gradient near the surface along with the depletion of the bulk solute molecules on the surface, for example, in a self-assembled monolayer adsorption kinetics. Thus, the analytical equations developed to calculate the collision at a fixed measuring frequency can be extended to map the whole curve over time. In the last section of this work, 3D diffusion simulations suggest that the analytical solution is valid to predict the adsorption rate of the bulk solute to a small group of adsorptive target molecules/area on a bouncing surface, which is a critical process in analyzing the kinetics of many bio-sensing platforms.</p> </div>

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“…A basic version of MATLAB 2014b with no toolbox is used for all simulations, i.e., single-CPU is used for all simulations. I use a previously coded fitting algorithm jcfit 40 (Github) to fit the curves. The two major random functions to generate the step motion of each molecule are from MATLAB, rand() creating evenly distributed random numbers and randn() generating Gaussian distributed random numbers.…”

Section: Methodsmentioning

confidence: 99%

“…For example, we have fitted kinetic data using our home-made code jcfit (Github) to optimizing simulations for a given set of data. 40,43 For the back-diffusion term, I am curious about simulating a routing experiment I have done before, self-assembly of alkanethiol molecules on gold or ZnO surface, 44 particularly in how long it takes to reach a whole surface coverage with no convection and stirring. Thus I simulate the adsorption kinetics of octadecanethiol in ethanol (concentration from 10 μM to 1 mM) on a flat surface assuming the adsorption probability in binary using the Metropolis Monte Carlo Method.…”

Section: The Surface Only When Empty Absorbs the Molecules Upon Collimentioning

confidence: 99%

Stochastic Adsorption of Diluted Solute Molecules at Interfaces

Chen

¹

2020

Preprint

Self Cite

0

View full text Add to dashboard Cite

<div> <p>Here an analytical solution of Fick’s 2<sup>nd</sup> law is used to predict the diffusion and the stochastic adsorption of single diluted solute molecules on flat and patterned surfaces. The equations are then compared to the results of several numerical Monte Carlo simulations using a random walk model. The 1D diffusion simulations clarify that the dependence of the solute-surface collision rate on the observation-time (measurement time resolution) is because of the multiple collisions of the same molecules over different time regions. It also surprisingly suggests that due to the self-mimetic fractal function of diffusion, the equation should be corrected by a factor of two. The absorption rate of solute on an adsorptive surface is found to follow a power-law decay function due to an evolving concentration gradient near the surface along with the depletion of the bulk solute molecules on the surface, for example, in a self-assembled monolayer adsorption kinetics. Thus, the analytical equations developed to calculate the collision at a fixed measuring frequency can be extended to map the whole curve over time. In the last section of this work, 3D diffusion simulations suggest that the analytical solution is valid to predict the adsorption rate of the bulk solute to a small group of adsorptive target molecules/area on a bouncing surface, which is a critical process in analyzing the kinetics of many bio-sensing platforms.</p> </div>

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A Two-Step Method for smFRET Data Analysis

Cited by 22 publications

References 34 publications

Simulations of camera-based single-molecule fluorescence experiments

Simulations of camera-based single-molecule fluorescence experiments

Stochastic Adsorption of Diluted Solute Molecules at Interfaces

Stochastic Adsorption of Diluted Solute Molecules at Interfaces

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