Accuracy of the detection of binding events using 3D single particle trackingReportar como inadecuado




Accuracy of the detection of binding events using 3D single particle tracking - Descarga este documento en PDF. Documentación en PDF para descargar gratis. Disponible también para leer online.

BMC Biophysics

, 10:3

Novel biophysical methods

Abstract

BackgroundNanoparticles can be used as markers to track the position of biomolecules, such as single proteins, inside living cells. The activity of a protein can sometimes be inferred from changes in the mobility of the attached particle. Mean Square Displacement analysis is the most common method to obtain mobility information from trajectories of tracked particles, such as the diffusion coefficient D. However, the precision of D sets a limit to discriminate changes in mobility caused by biological events from changes that reflect the stochasticity inherent to diffusion. This issue is of particular importance in an experiment aiming to quantify dynamic processes.

ResultsHere, we present simulations and 3D tracking experiments with Gold Nanorods freely diffusing in glycerol solution to establish the best analysis parameters to extract the diffusion coefficient. We applied this knowledge to the detection of a temporary change in diffusion, as it can occur due to the transient binding of a particle to an immobile structure within the cell, and tested its dependence on the magnitude of the change in diffusion and duration of this event.

ConclusionsThe simulations show that the spatial accuracy of particle tracking generally does not limit the detection of short binding events. Careful analysis of the magnitude of the change in diffusion and the number of frames per binding event is required for accurate quantification of such events.

KeywordsSingle particle tracking Diffusion Binding event Mean square displacement analysis AbbreviationsFCSFluorescence Correlation Spectroscopy

GNRGold nanorod

MSDMean Square Displacement

PEGPolyethilene-glycol

SMTSingle Molecule Tracking

SPTSingle Particle Tracking

TEMTransmission electron microscope

Electronic supplementary materialThe online version of this article doi:10.1186-s13628-017-0035-8 contains supplementary material, which is available to authorized users.

Download fulltext PDF



Autor: Sara Carozza - Jamie Culkin - John van Noort

Fuente: https://link.springer.com/







Documentos relacionados