Green fluorescent proteinThe green fluorescent protein (GFP) is a protein that exhibits bright green fluorescence when exposed to light in the blue to ultraviolet range. The label GFP traditionally refers to the protein first isolated from the jellyfish Aequorea victoria and is sometimes called avGFP. However, GFPs have been found in other organisms including corals, sea anemones, zoanithids, copepods and lancelets. The GFP from A. victoria has a major excitation peak at a wavelength of 395 nm and a minor one at 475 nm.
MicroscopyMicroscopy is the technical field of using microscopes to view objects and areas of objects that cannot be seen with the naked eye (objects that are not within the resolution range of the normal eye). There are three well-known branches of microscopy: optical, electron, and scanning probe microscopy, along with the emerging field of X-ray microscopy. Optical microscopy and electron microscopy involve the diffraction, reflection, or refraction of electromagnetic radiation/electron beams interacting with the specimen, and the collection of the scattered radiation or another signal in order to create an image.
Fluorescence microscopeA fluorescence microscope is an optical microscope that uses fluorescence instead of, or in addition to, scattering, reflection, and attenuation or absorption, to study the properties of organic or inorganic substances. "Fluorescence microscope" refers to any microscope that uses fluorescence to generate an image, whether it is a simple set up like an epifluorescence microscope or a more complicated design such as a confocal microscope, which uses optical sectioning to get better resolution of the fluorescence image.
FluorescenceFluorescence is the emission of light by a substance that has absorbed light or other electromagnetic radiation. It is a form of luminescence. In most cases, the emitted light has a longer wavelength, and therefore a lower photon energy, than the absorbed radiation. A perceptible example of fluorescence occurs when the absorbed radiation is in the ultraviolet region of the electromagnetic spectrum (invisible to the human eye), while the emitted light is in the visible region; this gives the fluorescent substance a distinct color that can only be seen when the substance has been exposed to UV light.
Optical microscopeThe optical microscope, also referred to as a light microscope, is a type of microscope that commonly uses visible light and a system of lenses to generate magnified images of small objects. Optical microscopes are the oldest design of microscope and were possibly invented in their present compound form in the 17th century. Basic optical microscopes can be very simple, although many complex designs aim to improve resolution and sample contrast. The object is placed on a stage and may be directly viewed through one or two eyepieces on the microscope.
Fluorescent tagIn molecular biology and biotechnology, a fluorescent tag, also known as a fluorescent label or fluorescent probe, is a molecule that is attached chemically to aid in the detection of a biomolecule such as a protein, antibody, or amino acid. Generally, fluorescent tagging, or labeling, uses a reactive derivative of a fluorescent molecule known as a fluorophore. The fluorophore selectively binds to a specific region or functional group on the target molecule and can be attached chemically or biologically.
Path tracingPath tracing is a computer graphics Monte Carlo method of rendering images of three-dimensional scenes such that the global illumination is faithful to reality. Fundamentally, the algorithm is integrating over all the illuminance arriving to a single point on the surface of an object. This illuminance is then reduced by a surface reflectance function (BRDF) to determine how much of it will go towards the viewpoint camera. This integration procedure is repeated for every pixel in the output image.
FluorophoreA 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).
ImmunofluorescenceImmunofluorescence is a technique used for light microscopy with a fluorescence microscope and is used primarily on biological samples. This technique uses the specificity of antibodies to their antigen to target fluorescent dyes to specific biomolecule targets within a cell, and therefore allows visualization of the distribution of the target molecule through the sample. The specific region an antibody recognizes on an antigen is called an epitope.
Confocal microscopyConfocal microscopy, most frequently confocal laser scanning microscopy (CLSM) or laser scanning confocal microscopy (LSCM), is an optical imaging technique for increasing optical resolution and contrast of a micrograph by means of using a spatial pinhole to block out-of-focus light in image formation. Capturing multiple two-dimensional images at different depths in a sample enables the reconstruction of three-dimensional structures (a process known as optical sectioning) within an object.
MicroscopeA microscope () is a laboratory instrument used to examine objects that are too small to be seen by the naked eye. Microscopy is the science of investigating small objects and structures using a microscope. Microscopic means being invisible to the eye unless aided by a microscope. There are many types of microscopes, and they may be grouped in different ways.
Distributed ray tracingDistributed ray tracing, also called distribution ray tracing and stochastic ray tracing, is a refinement of ray tracing that allows for the rendering of "soft" phenomena. Conventional ray tracing uses single rays to sample many different domains. For example, when the color of an object is calculated, ray tracing might send a single ray to each light source in the scene. This leads to sharp shadows, since there is no way for a light source to be partially occluded (another way of saying this is that all lights are point sources and have zero area).
Monte Carlo methodMonte Carlo methods, or Monte Carlo experiments, are a broad class of computational algorithms that rely on repeated random sampling to obtain numerical results. The underlying concept is to use randomness to solve problems that might be deterministic in principle. They are often used in physical and mathematical problems and are most useful when it is difficult or impossible to use other approaches. Monte Carlo methods are mainly used in three problem classes: optimization, numerical integration, and generating draws from a probability distribution.
Physically based renderingPhysically based rendering (PBR) is a computer graphics approach that seeks to render images in a way that models the lights and surfaces with optics in the real world. It is often referred to as "Physically Based Lighting" or "Physically Based Shading". Many PBR pipelines aim to achieve photorealism. Feasible and quick approximations of the bidirectional reflectance distribution function and rendering equation are of mathematical importance in this field. Photogrammetry may be used to help discover and encode accurate optical properties of materials.
StainingStaining is a technique used to enhance contrast in samples, generally at the microscopic level. Stains and dyes are frequently used in histology (microscopic study of biological tissues), in cytology (microscopic study of cells), and in the medical fields of histopathology, hematology, and cytopathology that focus on the study and diagnoses of diseases at the microscopic level. Stains may be used to define biological tissues (highlighting, for example, muscle fibers or connective tissue), cell populations (classifying different blood cells), or organelles within individual cells.
Optical sectioningOptical 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-dimensional reconstruction of a sample from images captured at different focal planes.
Geometrical opticsGeometrical optics, or ray optics, is a model of optics that describes light propagation in terms of rays. The ray in geometrical optics is an abstraction useful for approximating the paths along which light propagates under certain circumstances. The simplifying assumptions of geometrical optics include that light rays: propagate in straight-line paths as they travel in a homogeneous medium bend, and in particular circumstances may split in two, at the interface between two dissimilar media follow curved paths in a medium in which the refractive index changes may be absorbed or reflected.
Fluorescence in the life sciencesFluorescence is used in the life sciences generally as a non-destructive way of tracking or analysing biological molecules. Some proteins or small molecules in cells are naturally fluorescent, which is called intrinsic fluorescence or autofluorescence (such as NADH, tryptophan or endogenous chlorophyll, phycoerythrin or green fluorescent protein). Alternatively, specific or general proteins, nucleic acids, lipids or small molecules can be "labelled" with an extrinsic fluorophore, a fluorescent dye which can be a small molecule, protein or quantum dot.
Fluorescent lampA fluorescent lamp, or fluorescent tube, is a low-pressure mercury-vapor gas-discharge lamp that uses fluorescence to produce visible light. An electric current in the gas excites mercury vapor, which produces short-wave ultraviolet light that then causes a phosphor coating on the inside of the lamp to glow. A fluorescent lamp converts electrical energy into useful light much more efficiently than an incandescent lamp. The typical luminous efficacy of fluorescent lighting systems is 50–100 lumens per watt, several times the efficacy of incandescent bulbs with comparable light output.
Rendering (computer graphics)Rendering or image synthesis is the process of generating a photorealistic or non-photorealistic image from a 2D or 3D model by means of a computer program. The resulting image is referred to as the render. Multiple models can be defined in a scene file containing objects in a strictly defined language or data structure. The scene file contains geometry, viewpoint, texture, lighting, and shading information describing the virtual scene. The data contained in the scene file is then passed to a rendering program to be processed and output to a or raster graphics image file.
