Multiphoton Fluorescence Microscope
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Multiphoton Fluorescence Microscope
Two-photon excitation microscopy (TPEF or 2PEF) is a fluorescence imaging technique that allows imaging of living tissue up to about one millimeter in thickness, with 0.64 μm lateral and 3.35 μm axial spatial resolution. Unlike traditional fluorescence microscopy, in which the excitation wavelength is shorter than the emission wavelength, two-photon excitation requires simultaneous excitation by two photons with longer wavelength than the emitted light. Two-photon excitation microscopy typically uses near-infrared (NIR) excitation light which can also excite fluorescent dyes. However, for each excitation, two photons of NIR light are absorbed. Using infrared light minimizes scattering in the tissue. Due to the multiphoton absorption, the background signal is strongly suppressed. Both effects lead to an increased penetration depth for this technique. Two-photon excitation can be a superior alternative to confocal microscopy due to its deeper tissue penetration, efficient light d ...
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Isaac Abella
Isaac David Abella (June 20, 1934 – October 23, 2016) was a Canadian physicist who was a professor at the University of Chicago."Isaac Abella"
, Department of Physics, University of Chicago, 2008
He specialized in physics, quantum optics, and ."Marquis Who's Who in Science and Engineering"
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Pixel
In digital imaging, a pixel (abbreviated px), pel, or picture element is the smallest addressable element in a raster image, or the smallest point in an all points addressable display device. In most digital display devices, pixels are the smallest element that can be manipulated through software. Each pixel is a sample of an original image; more samples typically provide more accurate representations of the original. The intensity of each pixel is variable. In color imaging systems, a color is typically represented by three or four component intensities such as red, green, and blue, or cyan, magenta, yellow, and black. In some contexts (such as descriptions of camera sensors), ''pixel'' refers to a single scalar element of a multi-component representation (called a ''photosite'' in the camera sensor context, although ''sensel'' is sometimes used), while in yet other contexts (like MRI) it may refer to a set of component intensities for a spatial position. Etymology The w ...
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Photomultiplier
A photomultiplier is a device that converts incident photons into an electrical signal. Kinds of photomultiplier include: * Photomultiplier tube, a vacuum tube converting incident photons into an electric signal. Photomultiplier tubes (PMTs for short) are members of the class of vacuum tubes, and more specifically vacuum phototubes, which are extremely sensitive detectors of light in the ultraviolet, visible, and near-infrared ranges of the electromagnetic spectrum. ** Magnetic photomultiplier, developed by the Soviets in the 1930s. ** Electrostatic photomultiplier, a kind of photomultiplier tube demonstrated by Jan Rajchman of RCA Laboratories in Princeton, NJ in the late 1930s which became the standard for all future commercial photomultipliers. The first mass-produced photomultiplier, the Type 931, was of this design and is still commercially produced today. * Silicon photomultiplier, a solid-state device converting incident photons into an electric signal. Silicon photomultiplie ...
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Single-photon Avalanche Diode
A single-photon avalanche diode (SPAD) is a solid-state photodetector within the same family as photodiodes and avalanche photodiodes (APDs), while also being fundamentally linked with basic diode behaviours. As with photodiodes and APDs, a SPAD is based around a semi-conductor p-n junction that can be illuminated with ionizing radiation such as gamma, x-rays, beta and alpha particles along with a wide portion of the electromagnetic spectrum from ultraviolet (UV) through the visible wavelengths and into the infrared (IR). In a photodiode, with a low reverse bias voltage, the leakage current changes linearly with absorption of photons, i.e. the liberation of current carriers (electrons and/or holes) due to the internal photoelectric effect. However, in a SPAD, the reverse bias is so high that a phenomenon called impact ionisation occurs which is able to cause an avalanche current to develop. Simply, a photo-generated carrier is accelerated by the electric field in the device to ...
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Ti-sapphire Laser
Ti:sapphire lasers (also known as Ti:Al2O3 lasers, titanium-sapphire lasers, or Ti:sapphs) are tunable lasers which emit red and near-infrared light in the range from 650 to 1100 nanometers. These lasers are mainly used in scientific research because of their tunability and their ability to generate ultrashort pulses. Lasers based on Ti:sapphire were first constructed and invented in June 1982 by Peter Moulton at the MIT Lincoln Laboratory. Titanium-sapphire refers to the lasing medium, a crystal of sapphire (Al2O3) that is doped with Ti3+ ions. A Ti:sapphire laser is usually pumped with another laser with a wavelength of 514 to 532 nm, for which argon-ion lasers (514.5 nm) and frequency-doubled Nd:YAG, Nd:YLF, and Nd:YVO lasers (527-532 nm) are used. They are capable of laser operation from 670 nm to  nm wavelength. Ti:sapphire lasers operate most efficiently at wavelengths near 800 nm. Types of Ti:sapphire lasers Mode-locked oscillators Mo ...
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Pulsed Laser
Pulsed operation of lasers refers to any laser not classified as continuous wave, so that the optical power appears in pulses of some duration at some repetition rate. Silfvast, William T. (1996). ''Laser Fundamentals'', Cambridge University Press. This encompasses a wide range of technologies addressing a number of different motivations. Some lasers are pulsed simply because they cannot be run in continuous mode. In other cases the application requires the production of pulses having as large an energy as possible. Since the pulse energy is equal to the average power divided by the repetition rate, this goal can sometimes be satisfied by lowering the rate of pulses so that more energy can be built up in between pulses. In laser ablation for example, a small volume of material at the surface of a work piece can be evaporated if it is heated in a very short time, whereas supplying the energy gradually would allow for the heat to be absorbed into the bulk of the piece, never attaini ...
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Flux
Flux describes any effect that appears to pass or travel (whether it actually moves or not) through a surface or substance. Flux is a concept in applied mathematics and vector calculus which has many applications to physics. For transport phenomena, flux is a vector quantity, describing the magnitude and direction of the flow of a substance or property. In vector calculus flux is a scalar quantity, defined as the surface integral of the perpendicular component of a vector field over a surface. Terminology The word ''flux'' comes from Latin: ''fluxus'' means "flow", and ''fluere'' is "to flow". As ''fluxion'', this term was introduced into differential calculus by Isaac Newton. The concept of heat flux was a key contribution of Joseph Fourier, in the analysis of heat transfer phenomena. His seminal treatise ''Théorie analytique de la chaleur'' (''The Analytical Theory of Heat''), defines ''fluxion'' as a central quantity and proceeds to derive the now well-known express ...
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Quantum Yield
The quantum yield (Φ) of a radiation-induced process is the number of times a specific event occurs per photon absorbed by the system. Applications Fluorescence spectroscopy The fluorescence quantum yield is defined as the ratio of the number of photons emitted to the number of photons absorbed.Lakowicz, Joseph R. ''Principles of Fluorescence Spectroscopy'' (Kluwer Academic / Plenum Publishers 1999) p.10. Fluorescence quantum yield is measured on a scale from 0 to 1.0, but is often represented as a percentage. A quantum yield of 1.0 (100%) describes a process where each photon absorbed results in a photon emitted. Substances with the largest quantum yields, such as rhodamines, display the brightest emissions; however, compounds with quantum yields of 0.10 are still considered quite fluorescent. Quantum yield is defined by the fraction of excited state fluorophores that decay through fluorescence: where \Phi_ is the fluorescence quantum yield, k_ is the rate constant f ...
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Fluorophore
A fluorophore (or fluorochrome, similarly to a chromophore) is a fluorescent chemical compound that can re-emit light upon light excitation. Fluorophores typically contain several combined aromatic groups, or planar or cyclic molecules with several π bonds. Fluorophores are sometimes used alone, as a tracer in fluids, as a dye for staining of certain structures, as a substrate of enzymes, or as a probe or indicator (when its fluorescence is affected by environmental aspects such as polarity or ions). More generally they are covalently bonded to a macromolecule, serving as a marker (or dye, or tag, or reporter) for affine or bioactive reagents (antibodies, peptides, nucleic acids). Fluorophores are notably used to stain tissues, cells, or materials in a variety of analytical methods, i.e., fluorescent imaging and spectroscopy. Fluorescein, via its amine-reactive isothiocyanate derivative fluorescein isothiocyanate (FITC), has been one of the most popular fluorophores. Fro ...
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Optical Sectioning
Optical sectioning is the process by which a suitably designed microscope can produce clear images of focal planes deep within a thick sample. This is used to reduce the need for thin sectioning using instruments such as the microtome. Many different techniques for optical sectioning are used and several microscopy techniques are specifically designed to improve the quality of optical sectioning. Good optical sectioning, often referred to as good depth or z resolution, is popular in modern microscopy as it allows the three dimensions, three-dimensional reconstruction of a sample from images captured at different focal planes. Optical sectioning in traditional light microscopes In an ideal microscope, only light from the focal plane would be allowed to reach the detector (typically an observer or a Charge-coupled device, CCD) producing a clear image of the plane of the sample the microscope is focused on. Unfortunately a microscope is not this specific and light from sources outsi ...
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