Single-particle tracking

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Principle of single-particle tracking: The rectangles represent frames from an image acquisition at times t = 0, 1, 2, ... The tracked particles are represented as red circles, and in the last frame, the reconstructed trajectories are shown as blue lines Singleparticletracking.svg
Principle of single-particle tracking: The rectangles represent frames from an image acquisition at times t = 0, 1, 2, ... The tracked particles are represented as red circles, and in the last frame, the reconstructed trajectories are shown as blue lines

Single-particle tracking (SPT) is the observation of the motion of individual particles within a medium. The coordinates time series, which can be either in two dimensions (x, y) or in three dimensions (x, y, z), is referred to as a trajectory . The trajectory is typically analyzed using statistical methods to extract information about the underlying dynamics of the particle. [1] [2] [3] These dynamics can reveal information about the type of transport being observed (e.g., thermal or active), the medium where the particle is moving, and interactions with other particles. In the case of random motion, trajectory analysis can be used to measure the diffusion coefficient.

Contents

Applications

In life sciences, single-particle tracking is broadly used to quantify the dynamics of molecules/proteins in live cells (of bacteria, yeast, mammalian cells and live Drosophila embryos). [4] [5] [6] [7] [8] It has been extensively used to study the transcription factor dynamics in live cells. [9] [10] [11] This method has been extensively used in the last decade to understand the target-search mechanism of proteins in live cells. It addresses fundamental biological questions such as how a protein of interest finds its target in the complex cellular environment? how long does it take to find its target site for binding? what is the residence time of proteins binding to DNA? [5] Recently, SPT has been used to study the kinetics of protein translating and processing in vivo. For molecules which bind large structures such as ribosomes, SPT can be used to extract information about the binding kinetics. As ribosome binding increases the effective size of the smaller molecule, the diffusion rate decreases upon binding. By monitoring these changes in diffusion behavior, direct measurements of binding events are obtained. [12] [13] Furthermore, exogenous particles are employed as probes to assess the mechanical properties of the medium, a technique known as passive microrheology. [14] This technique has been applied to investigate the motion of lipids and proteins within membranes, [15] [16] molecules in the nucleus [8] and cytoplasm, [17] organelles and molecules therein, [18] lipid granules, [19] [20] [21] vesicles, and particles introduced in the cytoplasm or the nucleus. Additionally, single-particle tracking has been extensively used in the study of reconstituted lipid bilayers, [22] intermittent diffusion between 3D and either 2D (e.g., a membrane) [23] or 1D (e.g., a DNA polymer) phases, and synthetic entangled actin networks. [24] [25]

Methods

The most common type of particles used in single particle tracking are based either on scatterers, such as polystyrene beads or gold nanoparticles that can be tracked using bright field illumination, or fluorescent particles. For fluorescent tags, there are many different options with their own advantages and disadvantages, including quantum dots, fluorescent proteins, organic fluorophores, and cyanine dyes.

On a fundamental level, once the images are obtained, single-particle tracking is a two step process. First the particles are detected and then the localized different particles are connected in order to obtain individual trajectories.

Besides performing particle tracking in 2D, there are several imaging modalities for 3D particle tracking, including multifocal plane microscopy, [26] double helix point spread function microscopy, [27] and introducing astigmatism via a cylindrical lens or adaptive optics.

Brownian diffusion

See also

Related Research Articles

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In cell biology, the cytoplasm describes all material within a eukaryotic cell, enclosed by the cell membrane, except for the cell nucleus. The material inside the nucleus and contained within the nuclear membrane is termed the nucleoplasm. The main components of the cytoplasm are cytosol, the organelles, and various cytoplasmic inclusions. The cytoplasm is about 80% water and is usually colorless.

<span class="mw-page-title-main">Facilitated diffusion</span> Biological process

Facilitated diffusion is the process of spontaneous passive transport of molecules or ions across a biological membrane via specific transmembrane integral proteins. Being passive, facilitated transport does not directly require chemical energy from ATP hydrolysis in the transport step itself; rather, molecules and ions move down their concentration gradient reflecting its diffusive nature.

<span class="mw-page-title-main">Lipid bilayer</span> Membrane of two layers of lipid molecules

The lipid bilayer is a thin polar membrane made of two layers of lipid molecules. These membranes are flat sheets that form a continuous barrier around all cells. The cell membranes of almost all organisms and many viruses are made of a lipid bilayer, as are the nuclear membrane surrounding the cell nucleus, and membranes of the membrane-bound organelles in the cell. The lipid bilayer is the barrier that keeps ions, proteins and other molecules where they are needed and prevents them from diffusing into areas where they should not be. Lipid bilayers are ideally suited to this role, even though they are only a few nanometers in width, because they are impermeable to most water-soluble (hydrophilic) molecules. Bilayers are particularly impermeable to ions, which allows cells to regulate salt concentrations and pH by transporting ions across their membranes using proteins called ion pumps.

<span class="mw-page-title-main">Molecular dynamics</span> Computer simulations to discover and understand chemical properties

Molecular dynamics (MD) is a computer simulation method for analyzing the physical movements of atoms and molecules. The atoms and molecules are allowed to interact for a fixed period of time, giving a view of the dynamic "evolution" of the system. In the most common version, the trajectories of atoms and molecules are determined by numerically solving Newton's equations of motion for a system of interacting particles, where forces between the particles and their potential energies are often calculated using interatomic potentials or molecular mechanical force fields. The method is applied mostly in chemical physics, materials science, and biophysics.

<span class="mw-page-title-main">Fluid mosaic model</span> Describe the fluid mosaic model of plasma membrane

The fluid mosaic model explains various observations regarding the structure of functional cell membranes. According to this biological model, there is a lipid bilayer in which protein molecules are embedded. The phospholipid bilayer gives fluidity and elasticity to the membrane. Small amounts of carbohydrates are also found in the cell membrane. The biological model, which was devised by Seymour Jonathan Singer and Garth L. Nicolson in 1972, describes the cell membrane as a two-dimensional liquid that restricts the lateral diffusion of membrane components. Such domains are defined by the existence of regions within the membrane with special lipid and protein cocoon that promote the formation of lipid rafts or protein and glycoprotein complexes. In addition, it came before the other model that introduced the first bilayer. Another way to define membrane domains is the association of the lipid membrane with the cytoskeleton filaments and the extracellular matrix through membrane proteins. The current model describes important features relevant to many cellular processes, including: cell-cell signaling, apoptosis, cell division, membrane budding, and cell fusion. The fluid mosaic model is the most acceptable model of the plasma membrane. Its main function is to separate the contents of the cell from the exterior.

<span class="mw-page-title-main">Fluorescence recovery after photobleaching</span>

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<span class="mw-page-title-main">Lipid raft</span>

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<span class="mw-page-title-main">Single-molecule experiment</span>

A single-molecule experiment is an experiment that investigates the properties of individual molecules. Single-molecule studies may be contrasted with measurements on an ensemble or bulk collection of molecules, where the individual behavior of molecules cannot be distinguished, and only average characteristics can be measured. Since many measurement techniques in biology, chemistry, and physics are not sensitive enough to observe single molecules, single-molecule fluorescence techniques caused a lot of excitement, since these supplied many new details on the measured processes that were not accessible in the past. Indeed, since the 1990s, many techniques for probing individual molecules have been developed.

<span class="mw-page-title-main">Anomalous diffusion</span> Diffusion process with a non-linear relationship to time

Anomalous diffusion is a diffusion process with a non-linear relationship between the mean squared displacement (MSD), , and time. This behavior is in stark contrast to Brownian motion, the typical diffusion process described by Einstein and Smoluchowski, where the MSD is linear in time. Examples of anomalous diffusion in nature have been observed in biology in the cell nucleus, plasma membrane and cytoplasm.

<span class="mw-page-title-main">Molecular biophysics</span> Interdisciplinary research area

Molecular biophysics is a rapidly evolving interdisciplinary area of research that combines concepts in physics, chemistry, engineering, mathematics and biology. It seeks to understand biomolecular systems and explain biological function in terms of molecular structure, structural organization, and dynamic behaviour at various levels of complexity. This discipline covers topics such as the measurement of molecular forces, molecular associations, allosteric interactions, Brownian motion, and cable theory. Additional areas of study can be found on Outline of Biophysics. The discipline has required development of specialized equipment and procedures capable of imaging and manipulating minute living structures, as well as novel experimental approaches.

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<span class="mw-page-title-main">Lipid bilayer fusion</span>

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<span class="mw-page-title-main">Reduced dimensions form</span>

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<span class="mw-page-title-main">Two-state trajectory</span>

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<span class="mw-page-title-main">Interferometric scattering microscopy</span>

Interferometric scattering microscopy (iSCAT) refers to a class of methods that detect and image a subwavelength object by interfering the light scattered by it with a reference light field. The underlying physics is shared by other conventional interferometric methods such as phase contrast or differential interference contrast, or reflection interference microscopy. The key feature of iSCAT is the detection of elastic scattering from subwavelength particles, also known as Rayleigh scattering, in addition to reflected or transmission signals from supra-wavelength objects. Typically, the challenge is the detection of tiny signals on top of large and complex, speckle-like backgrounds. iSCAT has been used to investigate nanoparticles such as viruses, proteins, lipid vesicles, DNA, exosomes, metal nanoparticles, semiconductor quantum dots, charge carriers and single organic molecules without the need for a fluorescent label.

Rae Marie Robertson-Anderson is an American biophysicist who is Associate Professor at the University of San Diego. She works on soft matter physics and is particularly interested in the transport and molecular mechanics of biopolymer networks. Robertson-Anderson is a member of the Council on Undergraduate Research.

<span class="mw-page-title-main">Suliana Manley</span> American biophysicist

Suliana Manley is an American biophysicist. Her research focuses on the development of high-resolution optical instruments, and their application in studying the organization and dynamics of proteins. She is a professor at École Polytechnique Fédérale de Lausanne and heads the Laboratory of Experimental Biophysics.

Diego Krapf is an Argentine-Israeli-American physicist known for his work on anomalous diffusion and ergodicity breaking. He currently is a professor in the Department of Electrical and Computer Engineering at Colorado State University.

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