Fluorescence 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, 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. Fluorescent materials cease to glow nearly immediately when the radiation source stops, unlike phosphorescent materials, which continue to emit light for some time after.
Optical brighteners, optical brightening agents (OBAs), fluorescent brightening agents (FBAs), or fluorescent whitening agents (FWAs), are chemical compounds that absorb light in the ultraviolet and violet region of the electromagnetic spectrum, and re-emit light in the blue region through the phenomenon of fluorescence. These additives are often used to enhance the appearance of color of fabric and paper, causing a "whitening" effect; they make intrinsically yellow/orange materials look less so, by compensating the deficit in blue and purple light reflected by the material, with the blue and purple optical emission of the fluorophore.
The 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.
A blacklight, also called a UV-A light, Wood's lamp, or ultraviolet light, is a lamp that emits long-wave (UV-A) ultraviolet light and very little visible light. One type of lamp has a violet filter material, either on the bulb or in a separate glass filter in the lamp housing, which blocks most visible light and allows through UV, so the lamp has a dim violet glow when operating. Blacklight lamps which have this filter have a lighting industry designation that includes the letters "BLB". This stands for "blacklight blue". A second type of lamp produces ultraviolet but does not have the filter material, so it produces more visible light and has a blue color when operating. These tubes are made for use in "bug zapper" insect traps, and are identified by the industry designation "BL". This stands for "blacklight".
Staining is a technique used to enhance contrast in samples, generally at the microscopic level. Stains and dyes are frequently used in histology, in cytology, 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, cell populations, or organelles within individual cells.
A fluorophore 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.
Congo red is an organic compound, the sodium salt of 3,3′-([1,1′-biphenyl]-4,4′-diyl)bis(4-aminonaphthalene-1-sulfonic acid). It is an azo dye. Congo red is water-soluble, yielding a red colloidal solution; its solubility is greater in organic solvents. However, the use of Congo red has long been abandoned, primarily because of its carcinogenic properties.
A 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.
DAPI, or 4′,6-diamidino-2-phenylindole, is a fluorescent stain that binds strongly to adenine–thymine-rich regions in DNA. It is used extensively in fluorescence microscopy. As DAPI can pass through an intact cell membrane, it can be used to stain both live and fixed cells, though it passes through the membrane less efficiently in live cells and therefore provides a marker for membrane viability.
Hoechst stains are part of a family of blue fluorescent dyes used to stain DNA. These bis-benzimides were originally developed by Hoechst AG, which numbered all their compounds so that the dye Hoechst 33342 is the 33,342nd compound made by the company. There are three related Hoechst stains: Hoechst 33258, Hoechst 33342, and Hoechst 34580. The dyes Hoechst 33258 and Hoechst 33342 are the ones most commonly used and they have similar excitation–emission spectra.
Nile blue is a stain used in biology and histology. It may be used with live or fixed cells, and imparts a blue colour to cell nuclei.
Nile red is a lipophilic stain. Nile red stains intracellular lipid droplets yellow. In most polar solvents, Nile red will not fluoresce; however, when in a lipid-rich environment, it can be intensely fluorescent, with varying colors from deep red to strong yellow-gold emission. The dye is highly solvatochromic and its emission and excitation wavelength both shift depending on solvent polarity and in polar media will hardly fluoresce at all.
Propidium iodide is a fluorescent intercalating agent that can be used to stain cells and nucleic acids. PI binds to DNA by intercalating between the bases with little or no sequence preference. When in an aqueous solution, PI has a fluorescent excitation maximum of 493 nm (blue-green), and an emission maximum of 636 nm (red). After binding DNA, the quantum yield of PI is enhanced 20-30 fold, and the excitation/emission maximum of PI is shifted to 535 nm (green) / 617 nm (orange-red). Propidium iodide is used as a DNA stain in flow cytometry to evaluate cell viability or DNA content in cell cycle analysis, or in microscopy to visualize the nucleus and other DNA-containing organelles. Propidium Iodide is not membrane-permeable, making it useful to differentiate necrotic, apoptotic and healthy cells based on membrane integrity. PI also binds to RNA, necessitating treatment with nucleases to distinguish between RNA and DNA staining. PI is widely used in fluorescence staining and visualization of the plant cell wall.
Periodic acid–Schiff (PAS) is a staining method used to detect polysaccharides such as glycogen, and mucosubstances such as glycoproteins, glycolipids and mucins in tissues. The reaction of periodic acid oxidizes the vicinal diols in these sugars, usually breaking up the bond between two adjacent carbons not involved in the glycosidic linkage or ring closure in the ring of the monosaccharide units that are parts of the long polysaccharides, and creating a pair of aldehydes at the two free tips of each broken monosaccharide ring. The oxidation condition has to be sufficiently regulated so as to not oxidize the aldehydes further. These aldehydes then react with the Schiff reagent to give a purple-magenta color. A suitable basic stain is often used as a counterstain.
Acridine orange is an organic compound that serves as a nucleic acid-selective fluorescent dye with cationic properties useful for cell cycle determination. Acridine orange is cell-permeable, which allows the dye to interact with DNA by intercalation, or RNA via electrostatic attractions. When bound to DNA, acridine orange is very similar spectrally to an organic compound known as fluorescein. Acridine orange and fluorescein have a maximum excitation at 502nm and 525 nm (green). When acridine orange associates with RNA, the fluorescent dye experiences a maximum excitation shift from 525 nm (green) to 460 nm (blue). The shift in maximum excitation also produces a maximum emission of 650 nm (red). Acridine orange is able to withstand low pH environments, allowing the fluorescent dye to penetrate acidic organelles such as lysosomes and phagolysosomes that are membrane-bound organelles essential for acid hydrolysis or for producing products of phagocytosis of apoptotic cells. Acridine orange is used in epifluorescence microscopy and flow cytometry. The ability to penetrate the cell membranes of acidic organelles and cationic properties of acridine orange allows the dye to differentiate between various types of cells. The shift in maximum excitation and emission wavelengths provides a foundation to predict the wavelength at which the cells will stain.
The KOH Test for Candida albicans, also known as a potassium hydroxide preparation or KOH prep, is a quick, inexpensive fungal test to differentiate dermatophytes and Candida albicans symptoms from other skin disorders like psoriasis and eczema.
The DyLight Fluor family of fluorescent dyes are produced by Dyomics in collaboration with Thermo Fisher Scientific. DyLight dyes are typically used in biotechnology and research applications as biomolecule, cell and tissue labels for fluorescence microscopy, cell biology or molecular biology.
Fluorescence 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. 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. Several techniques exist to exploit additional properties of fluorophores, such as fluorescence resonance energy transfer, where the energy is passed non-radiatively to a particular neighbouring dye, allowing proximity or protein activation to be detected; another is the change in properties, such as intensity, of certain dyes depending on their environment allowing their use in structural studies.
Cryptomycota , Rozellida, or Rozellomycota are a clade of micro-organisms that are either fungi or a sister group to fungi. They differ from classical fungi in that they lack chitinous cell walls at any trophic stage in their lifecycle, as reported by Jones and colleagues in 2011. Despite their unconventional feeding habits, chitin has been observed in the inner layer of resting spores, and in immature resting spores for some species of Rozella, as indicated with calcofluor-white stain as well as the presence of a fungal-specific chitin synthase gene.
Pacific Blue, or systematically 3-carboxy-6,8-difluoro-7-hydroxycoumarin, is a fluorophore used in cell biology. Its excitation maximum lies at 401 nm, while its emission maximum is at 452 nm. In contrast to the less acidic 7-hydroxy-3-carboxycoumarin (pKa=7.0), the high acidity of the phenol of Pacific Blue (pKa=3.7) causes its fluorescence to remain very high at neutral pH.
