 | We've compiled all of the key terms we use throughout Molecular Probes School of Fluorescence in one location. As we expand the School of Fluorescence, we will continue to add key terms to this glossary. Browse the glossary to find the definition for the term you're interested in. |
| Antigen | A material that causes an organism to have an immune response and generate antibodies |
| Background fluorescence | Nonspecific and unwanted fluorescence produced by a sample, vessel, or imaging medium, or from fluorophores not bound to specific targets. | |
| Bandpass filter | An optical filter through which only light of a given wavelength range can pass. All light with a higher or lower wavelength is blocked. Normally identified by the middle-value wavelength and the width of a band. | |
| Binning | Combining a group of pixels into a single pixel to reduce the total number of pixels in an image. This can reduce image noise and shorten the necessary exposures times but can also lower resolution. | |
| Blebbing | Portions of the cell membrane that appear as protrusions; these eventually separate from a cell during cell death. | |
| Bleed-through | The incorrect detection of emitted light from a fluorophore in a channel intended for another fluorophore. | |
| Blocking solution | A solution used to prevent non-specific, low-affinity binding in ICC samples. | |
| Brightfield microscope | A light microscope in which the sample is illuminated with white light and images are formed due to the light-absorbing properties of structures within a sample. |
| CCD camera | Charge-coupled device; an image sensor that converts light into electrons to create digital images. | |
| Cell-permeant | When a compound is able to freely diffuse across the plasma membrane, into and out of a cell. | |
| Cellular membrane | Phospholipid bilayer that forms the outer barrier of a cell or encloses various compartments within a cell. | |
| Crosslinking | A bond that links proteins with other nearby nitrogen atoms in protein or DNA through a -CH2- linkage. | |
| Cytoskeleton | The network of filaments and tubules that provides a cell's structure and shape. |
| Detector | The part of a fluorescence imaging system that detects the signal from a sample. This could be the oculars on the microscope directing light to your eyes or an electrical component such as a camera that records photons and creates digital images. | |
| Dichroic beamsplitter (filter) | An optical device that splits a beam of light, reflecting the shorter wavelengths (a defined range) and allowing all other higher wavelengths to pass through. | |
| Dynamic range | The range of light intensities detectable by a camera that occurs between background noise and pixel saturation. |
| Emission | Photons emitted from an object as excited fluorescent molecules relax to a lower-energy state. | |
| Emission filter | An optical device that limits the emitted light collected from a sample to a defined set of wavelengths. | |
| Epifluorescence | When illumination light and emission light travel through the same objective lens. | |
| Epifluorescence illumination | The most common arrangement of optical components in a fluorescence microscope in which excitation and emitted light both pass through the same objective lens above the sample. | |
| Epithelial | A type of animal cell that lines the internal and external surfaces of the body. | |
| Epitope | The specific area on an antigen that an antibody binds to. | |
| Excitation filter | An optical device that limits the excitation light applied to a sample to a defined wavelength range. | |
| Excite | Driving an electron in a fluorophore to a higher energy state as a photon of light is absorbed. | |
| Exposure time | The length of time a camera is exposed to light emitted from a sample while capturing an image. | |
| Extinction coefficient | The capacity of a fluorophore to absorb light at a given wavelength in vitro. |
| Fibroblasts | A type of animal cell typically found in connective tissue. | |
| Fluorescence | The emission of light from a sample that occurs as excited fluorescent molecules relax to a lower energy state. | |
| Fluorescence microscope | A light microscope that creates images based on the emission of fluorescent light from a sample. | |
| Fluorescent dye | Also called fluorochrome; a fluorescent chemical compound that can re-emit light following light excitation. | |
| Fluorescent proteins | A class of proteins that contain amino acid sections that can act as fluorophores. | |
| Fluorophore | A fluorescent chemical compound that can re-emit light following light excitation. | |
| Fluorophore filter set | A selection of excitation, emission, and dichroic filters designed for detecting light from a given fluorophore. | |
| Fusion protein | A protein formed by the expression of an artificial gene formed by linking two or more different genes or gene fragments. |
| Grayscale | An image composed of shades of gray ranging from white to black. |
| HBSS | Hank's Balanced Salt Solution; a saline-based solution that has a physiological pH and salt concentration. | |
| Heat-inactivated serum | Serum that has been heated to 56°C for 30–60 minutes to remove any enzymatic or other protein activity normally present. |
| Immunocytochemistry | Abbreviated as ICC. In this technique, a primary antibody is used to localize a specific antigen or protein (target) in a cell. The primary antibody specifically binds to the target and is usually visualized microscopically by binding of a secondary antibody that has a fluorescent label. | |
| Immunofluorescence | The use of antibodies labeled with fluorescent dyes to detect a specific antigen or protein in a cell. | |
| Immunofluorescent labeling | A technique that uses fluorescence-labeled antibodies to detect target antigens in order to visualize specific biomarkers in a cell. | |
| Immunolabeling | The technique for detecting a specific antigen in a sample using a fluorescently-labeled antibody. | |
| Intensity | A measure of the quantity of light per given area of sample. | |
| Inverted microscope | A microscope with objectives placed below the sample stage; samples viewed on this type of microscope will have to be inverted on the stage. |
| Laser scanning confocal microscope | A compound light microscope that uses lasers as excitation sources and pinholes to limit the detected light to one focal plane. | |
| Light | The range of electromagnetic radiation visible to the human eye. | |
| Light path | The sequence of mirrors, lenses, and filters through which light passes from the excitation source to the detector. | |
| Light source | A device such as a lamp or laser used to illuminate a sample. | |
| Lipid transfection | Using lipid micelles to insert genetic material in the form of DNA or RNA into mammalian cells | |
| Long working distance | An objective lens that can provide a good image with a range of coverglass thicknesses. | |
| Longpass filter | An optical filter through which only light above a given wavelength range can pass. All light with a lower wavelength is blocked. |
| Magnification | The visual enlargement of a sample, viewed with a microscope. | |
| Membrane potential | The difference in electrical potential across a cellular membrane such as the plasma membrane or mitochondrial membrane. | |
| Metabolic | The collection of biochemical reactions occurring in a cell. | |
| Mitochondria | Organelles in eukaryotic cells responsible for generating ATP. | |
| Mounting medium | Any medium in which the sample is bathed during imaging. |
| Noise | Nonspecific and undesired fluorescence produced by components of a microscope, including the excitation source, camera, and external light source. | |
| Non-specific binding | Non-covalent binding that is not specific to a particular target, ligand, or epitope. It is thought to be due to "stickiness", or hydrophobic interactions and can be minimized with additional washes and/or by adding competing hydrophobic molecules. | |
| Numerical aperture | A measurement of the ability of an objective to gather light and provide a high-resolution image. |
| Osmolarity | Also referred to as osmotic concentration; it is the number of osmoles of solute per liter of solution (Osm/L). |
| PBS | Phosphate-buffered saline. A solution containing water, sodium chloride, sodium phosphate, and in some cases, potassium chloride and potassium phosphate. The ions and osmolarity are matched to those found in the human body. | |
| Photobleach curve | An exponential decay curve that displays the reduction in fluorescence as a fluorophore is continuously exposed to light. | |
| Photobleaching | Destruction of an excited fluorophore due to photosensitized generation of reactive oxygen species. | |
| Photon | The smallest discrete unit of light. | |
| Photostability | The ability of a fluorophore to resist signal degradation from photobleaching. | |
| Phototoxicity | Damaging effects on live cells caused by light or by the generation of reactive oxygen species that follows the illumination of a fluorophore. | |
| Pixel | The smallest discrete area of an image. | |
| Plasma membrane | The membrane surrounding an animal cell which is composed of a phospholipid bilayer and proteins. | |
| Primary antibody | An antibody generated to specifically recognize (bind to) a target of interest (targets can be proteins, peptides, carbohydrates, phosphorylation sites on proteins, or other small molecules). A good primary antibody will be both high affinity (binds tightly to the target) and specific (doesn't bind tightly to any other molecule). | |
| Pseudocolor | Applying color to a given pixel in an image, usually the color will match the emission color of the fluorophore. |
| Quantum yield | A measure of efficiency of a fluorophore, defined as the fraction of absorbed photons that result in emitted fluorescence in vitro. |
| Refractive index | A measure of how much light is bent as it passes through two objects of different composition. | |
| Resolution | The shortest distance between two points on a sample that can still be distinguished as separate. | |
| Ringer's solution | A clear, buffered, salt solution that is isotonic and has a physiologic pH. |
| Sample | The object to be imaged; typically a biological sample such as tissue or cells. | |
| Saturated | A condition when a given pixel in a camera has collected the maximum number of photons. Additional detected photons will not be counted beyond the saturation point. | |
| Secondary antibody | An antibody generated to recognize (bind to) other antibodies, typically labeled with a probe that makes the complex visible under microscopy Non-covalent binding involving hydrogen bonds, van der Waals forces, and ionic bonds that is specific for an antibody and a target or epitope. | |
| Specific binding | Specifically bound molecules cannot be easily washed away with PBS or blocking buffer. | |
| Stokes shift | The difference in wavelength between the excitation and emission maxima of a fluorophore. | |
| Subcellular | Structures such as organelles that are smaller than, and generally contained within, a cell. |
| Transduction | The process of inserting genetic material into a mammalian cell using a viral vector. | |
| Transfected | Mammalian cells that contain inserted genetic material (DNA or RNA). |
| Upright microscope | A microscope with objectives placed above the sample stage; samples viewed on this type of microscope will have to be placed upright on the stage. |
| v/v | Volume percent concentration, v/v% = [(volume of solute)/(volume of solution)] x 100. For example, a 5% v/v solution would contain 5 mL of solute diluted to a final volume of 100 mL. | |
| Viability assay | One or more fluorescent reagents used to determine the health of cells in a sample. |
| w/v | Weight/volume percent, w/v% = [(weight of solute (in g))/(volume of solvent (in mL))] x 100. For example a 5% w/v solution would contain 5 g of solute in 100 mL of solvent. | |
| Washing | A rinse after incubations to dilute or remove the non-specifically bound original material, generally performed using PBS. | |
| Wavelength | The distance between peaks of two consecutive light waves. |
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