Total Internal Reflection Microscopy
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Total Internal Reflection Microscopy
Total internal reflection microscopy is a specialized optical imaging technique for object tracking and detection utilizing the light scattered from an evanescent field in the vicinity of a dielectric interface. Its advantages are a high signal-to-noise ratio and a high spatial resolution in the vertical dimension. Background Total internal reflection of light occurs at the interface between materials of differing indices of refraction at incident angles greater than the critical angle, \theta_c, where :\theta_c = \sin^(n_2/n_1) and n_1 is the index of the incident medium and n_2 the index of the transmission medium and \theta_c is measured from the normal to the interface. Under conditions of total internal reflection, the electromagnetic field in the transmission medium takes on the form of an evanescent wave, whose intensity I(z) decays exponentially with distance from the interface such that, :I(z) = I_0 e^ with \beta = \frac\sqrt. For practical purposes, the transmission ...
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Total Internal Reflection Scattering
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Evanescent Field
In electromagnetics, an evanescent field, or evanescent wave, is an oscillating electric and/or magnetic field that does not propagate as an electromagnetic wave but whose energy is spatially concentrated in the vicinity of the source (oscillating charges and currents). Even when there is a propagating electromagnetic wave produced (e.g., by a transmitting antenna), one can still identify as an evanescent field the component of the electric or magnetic field that cannot be attributed to the propagating wave observed at a distance of many wavelengths (such as the far field of a transmitting antenna). A hallmark of an evanescent field is that there is no net energy flow in that region. Since the net flow of electromagnetic energy is given by the average Poynting vector, this means that the Poynting vector in these regions, as averaged over a complete oscillation cycle, is zero. Use of the term In many cases one cannot simply say that a field is or is not "evanescent": Having Po ...
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Dielectric
In electromagnetism, a dielectric (or dielectric medium) is an electrical insulator that can be polarised by an applied electric field. When a dielectric material is placed in an electric field, electric charges do not flow through the material as they do in an electrical conductor, because they have no loosely bound, or free, electrons that may drift through the material, but instead they shift, only slightly, from their average equilibrium positions, causing dielectric polarisation. Because of dielectric polarisation, positive charges are displaced in the direction of the field and negative charges shift in the direction opposite to the field (for example, if the field is moving parallel to the positive ''x'' axis, the negative charges will shift in the negative ''x'' direction). This creates an internal electric field that reduces the overall field within the dielectric itself. If a dielectric is composed of weakly bonded molecules, those molecules not only become polaris ...
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Signal-to-noise Ratio
Signal-to-noise ratio (SNR or S/N) is a measure used in science and engineering that compares the level of a desired signal to the level of background noise. SNR is defined as the ratio of signal power to the noise power, often expressed in decibels. A ratio higher than 1:1 (greater than 0 dB) indicates more signal than noise. SNR, bandwidth, and channel capacity of a communication channel are connected by the Shannon–Hartley theorem. Definition Signal-to-noise ratio is defined as the ratio of the power of a signal (meaningful input) to the power of background noise (meaningless or unwanted input): : \mathrm = \frac, where is average power. Both signal and noise power must be measured at the same or equivalent points in a system, and within the same system bandwidth. Depending on whether the signal is a constant () or a random variable (), the signal-to-noise ratio for random noise becomes: : \mathrm = \frac where E refers to the expected value, i.e. in this ...
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Total Internal Reflection
Total internal reflection (TIR) is the optical phenomenon in which waves arriving at the interface (boundary) from one medium to another (e.g., from water to air) are not refracted into the second ("external") medium, but completely reflected back into the first ("internal") medium. It occurs when the second medium has a higher wave speed (i.e., lower refractive index) than the first, and the waves are incident at a sufficiently oblique angle on the interface. For example, the water-to-air surface in a typical fish tank, when viewed obliquely from below, reflects the underwater scene like a mirror with no loss of brightness (Fig.1). TIR occurs not only with electromagnetic waves such as light and microwaves, but also with other types of waves, including sound and water waves. If the waves are capable of forming a narrow beam (Fig.2), the reflection tends to be described in terms of " rays" rather than waves; in a medium whose properties are independent of direction, such ...
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Evanescent Wave
In electromagnetics, an evanescent field, or evanescent wave, is an oscillating electric and/or magnetic field that does not propagate as an electromagnetic wave but whose energy is spatially concentrated in the vicinity of the source (oscillating charges and currents). Even when there is a propagating electromagnetic wave produced (e.g., by a transmitting antenna), one can still identify as an evanescent field the component of the electric or magnetic field that cannot be attributed to the propagating wave observed at a distance of many wavelengths (such as the far field of a transmitting antenna). A hallmark of an evanescent field is that there is no net energy flow in that region. Since the net flow of electromagnetic energy is given by the average Poynting vector, this means that the Poynting vector in these regions, as averaged over a complete oscillation cycle, is zero. Use of the term In many cases one cannot simply say that a field is or is not "evanescent": Having P ...
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Total Internal Reflection Fluorescence Microscopy
A total internal reflection fluorescence microscope (TIRFM) is a type of microscope with which a thin region of a specimen, usually less than 200 nanometers can be observed. TIRFM is an imaging modality which uses the excitation of fluorescent cells in a thin optical specimen section that is supported on a glass slide. The technique is based on the principle that when excitation light is totally internally reflected in a transparent solid coverglass at its interface with a liquid medium, an electromagnetic field, also known as an evanescent wave, is generated at the solid-liquid interface with the same frequency as the excitation light. The intensity of the evanescent wave exponentially decays with distance from the surface of the solid so that only fluorescent molecules within a few hundred nanometers of the solid are efficiently excited. Two-dimensional images of the fluorescence can then be obtained, although there are also mechanisms in which three-dimensional information on t ...
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Maxwell–Boltzmann Distribution
In physics (in particular in statistical mechanics), the Maxwell–Boltzmann distribution, or Maxwell(ian) distribution, is a particular probability distribution named after James Clerk Maxwell and Ludwig Boltzmann. It was first defined and used for describing particle speeds in idealized gases, where the particles move freely inside a stationary container without interacting with one another, except for very brief collisions in which they exchange energy and momentum with each other or with their thermal environment. The term "particle" in this context refers to gaseous particles only ( atoms or molecules), and the system of particles is assumed to have reached thermodynamic equilibrium.''Statistical Physics'' (2nd Edition), F. Mandl, Manchester Physics, John Wiley & Sons, 2008, The energies of such particles follow what is known as Maxwell–Boltzmann statistics, and the statistical distribution of speeds is derived by equating particle energies with kinetic energy. Mathema ...
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Partition Function (statistical Mechanics)
In physics, a partition function describes the statistical properties of a system in thermodynamic equilibrium. Partition functions are functions of the thermodynamic state variables, such as the temperature and volume. Most of the aggregate thermodynamic variables of the system, such as the total energy, free energy, entropy, and pressure, can be expressed in terms of the partition function or its derivatives. The partition function is dimensionless. Each partition function is constructed to represent a particular statistical ensemble (which, in turn, corresponds to a particular free energy). The most common statistical ensembles have named partition functions. The canonical partition function applies to a canonical ensemble, in which the system is allowed to exchange heat with the environment at fixed temperature, volume, and number of particles. The grand canonical partition function applies to a grand canonical ensemble, in which the system can exchange both heat and ...
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Boltzmann Constant
The Boltzmann constant ( or ) is the proportionality factor that relates the average relative kinetic energy of particles in a gas with the thermodynamic temperature of the gas. It occurs in the definitions of the kelvin and the gas constant, and in Planck's law of black-body radiation and Boltzmann's entropy formula, and is used in calculating thermal noise in resistors. The Boltzmann constant has dimensions of energy divided by temperature, the same as entropy. It is named after the Austrian scientist Ludwig Boltzmann. As part of the 2019 redefinition of SI base units, the Boltzmann constant is one of the seven " defining constants" that have been given exact definitions. They are used in various combinations to define the seven SI base units. The Boltzmann constant is defined to be exactly . Roles of the Boltzmann constant Macroscopically, the ideal gas law states that, for an ideal gas, the product of pressure and volume is proportional to the product of amount ...
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Single Particle Tracking
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. 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. 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). It has been extensively used to study the transcription factor dynamics in live cells. This method h ...
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Diffusion
Diffusion is the net movement of anything (for example, atoms, ions, molecules, energy) generally from a region of higher concentration to a region of lower concentration. Diffusion is driven by a gradient in Gibbs free energy or chemical potential. It is possible to diffuse "uphill" from a region of lower concentration to a region of higher concentration, like in spinodal decomposition. The concept of diffusion is widely used in many fields, including physics ( particle diffusion), chemistry, biology, sociology, economics, and finance (diffusion of people, ideas, and price values). The central idea of diffusion, however, is common to all of these: a substance or collection undergoing diffusion spreads out from a point or location at which there is a higher concentration of that substance or collection. A gradient is the change in the value of a quantity, for example, concentration, pressure, or temperature with the change in another variable, usually distance. ...
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