Glossary of Optical Terms

An active crystal device that works by setting up radio-frequency acoustical vibrations in the crystal and creating, in effect, a bulk transmission diffraction grating. By varying the frequency, one can rapidly tune the filter to diffract out a desired wavelength of light and transmit this wavelength out of the device.

The angle between the optical axis of the light incident on the surface of a filter and the axis normal to this surface.

A shift in the direction of a light beam from the true optical axis of the system, measured in units of angle such as arc-minutes (1/60 of a degree) or arc-seconds (1/60 of an arc-minute).

An adjustable diaphragm located in the illumination optics controls the numerical aperture of the illuminating beam and affects the brightness of the beam.

Also Blocking level. A measure of the out-of-band attenuation of an optical filter over an extended range of the spectrum. The attenuation level is often defined in units of optical density (see optical density).

In fluorescence microscopy, any fluorescence from substances other than fluorochromes, including primary fluorescence from the specimen, fluorescence from immersion oils and other media, and fluorescence from glass optical components within the microscope.

The average calculated over the useful transmission region of a filter rather than over the entire spectrum. For a bandpass filter, this region spans the FWHM of the transmission band.

Any detectable light that is not a desired primary or indirect fluorescent emission. Sources of background include cross-talk between excitation and emission filters, light leaking through pinholes in filters, and electronic noise in cameras, as well as autofluorescence.

An optical filter that has a well-defined short wavelength cut-on and long wavelength cut-off. Bandpass filters are denoted by their center wavelength and bandwidth.

Also FWHM (full width at half of maximum transmission). For optical bandpass filters, typically the separation between the cut-on and cut-off wavelengths at 50% of peak transmission, though sometimes calculated as a bandwidth at 10% of peak transmission is specified, as an example.

See Emission filter.

A thin-film interference coating that is incorporated into a bandpass interference filter to extend the blocking range of the primary coating in the filter. A blocker is usually a very wide-band bandpass filter having high transmission in the band of the primary filter.

See Attenuation level.

The range of wavelengths over which an optical filter maintains a specified attenuation level.

A kind of diascopic illumination in which the field of view is illuminated directly. Also, the type of condenser used for this kind of illumination.

For optical bandpass filters, the arithmetic mean of the cut-on and cut-off wavelengths at 50% of peak transmission.

The surface area of an optical filter which is free of any defects or obstructions. With interference filters the clear aperture is often delimited by an annulus of metal or opaque material.

A type of illumination optics in which the image of the light source is focused onto the specimen plane, in contrast to Köhler illumination optics. See also Köhler illumination.

The minimum attenuation level (over a specified wavelength range) of two filters placed together in series. The transmission spectrum of the beamsplitter is sometimes included in this evaluation.

The edge produced when transitioning between transmission and blocking. The point at which a filter transmits 50% and longer wavelengths are blocked.

The edge produced when transitioning between blocking and transmission. The point at which a filter transmits 50% and longer wavelengths are transmitted.

A kind of diascopic illumination in which the specimen is illuminated obliquely, i.e., at angles that will not enter the objective directly. Also, the type of condenser used for this kind of illumination.

Illumination uses light transmitted through the specimen, using a condenser to focus the light.

Also Dichroic mirror or Dichromatic beamsplitter. A special mirror housed in the filter cube that selectively reflects the excitation light filtered by the exciter and transmits the emitted fluorescence. Dichroic beamsplitters can also be found in any other part of a microscope system where light needs to be split or combined by wavelength.

Another term for either a shortpass or longpass optical filter. The term usually denotes a filter with a very sharp cut-on or cut-off.

Also Barrier filter or Emitter. A color filter that attenuates all of the light transmitted by the excitation filter and very efficiently transmits any fluorescence emitted by the specimen.

See Emission filter.

A fluorescence microscope that illuminates the specimen episcopically (i.e., with light reflected onto the specimen).

Also Incident-light illumination. Illumination with light reflected onto the specimen rather than transmitted through the specimen. The illuminating light is reflected through the objective by means of a beamsplitter that is either partially reflective or a dichroic.

See Fluorescence.

Also Exciter. A color filter that transmits only those wavelengths of the illumination light that efficiently excites a specific dye or dyes. See Emission filter.

See Excitation filter.

A measure of the absorption characteristics of a fluorochrome.

See Photobleaching.

An adjustable diaphragm located in the illumination optics which controls the area of illumination on the specimen.

See Filter cube.

A removable cube-shaped unit that holds a matched set of fluorescence filters. This set usually includes an excitation filter and emission filter, but always includes a dichroic beamsplitter.

Also Absorption glass. Colored glass that is manufactured for technical and scientific applications. The most common types of filter glass used in fluorescence microscopy are UV-transmitting “black glass” filters; IR-absorbing heat filters; and yellow, orange, and red sharp-cut longpass filters. Filter glass selectively attenuates light by absorption, so the spectral performance is dependent on the thickness of the glass. The blocking properties of filter glass are also independent of the angle-of-incidence of the incoming light.

A molecular phenomenon in which a substance absorbs light (as a photon), then radiates part of this absorbed energy as light of another color, one of lower energy and thus longer wavelength. This process is known as excitation and emission. Fluorescence is distinguishable from other types of luminescence in that the process of excitation and emission occurs nearly instantaneously (i.e., on the order of nanoseconds).

Also Fluorophore. A fluorochrome that has been conjugated to an active substance, such as a protein, antibody, or nucleic acid, in order to selectively stain a targeted substance within the specimen.

A fluorescent dye used either directly as a specimen stain or conjugated to an active substance to make a fluorescent probe.

See Fluorescent probe.

The side of a beamsplitter that is designed to face the incident light. In a filter cube, this is the side that faces both the light source and the specimen. Beamsplitters generally perform better when oriented correctly.

See Bandwidth.

A measure of the wavelength-dependent time delay introduced by phase shifts and optical path length differences upon reflection from, or transmission through, a thin-film interference coating or other optical component. The usual unit of measure for GDD is fsec^2.  The GDD of a coating may be an important consideration when using ultra-fast pulsed lasers.

The angle between the most oblique ray of a convergent or divergent light beam and the optical axis of the beam. See also Numerical aperture.

An optical filter that attenuates infrared radiation, but transmits the visible. Attenuation is achieved by means of absorption (using filter glass), reflection (using a thin-film interference coating, often called a hot mirror), or a combination of the two.

See Heat filter.

Also Secondary fluorescence. In fluorescence microscopy, fluorescence emitted by fluorochromes introduced into a specimen as a stain or probe. See Primary fluorescence.

An optical configuration employed by some microscopes in which the objective forms an image at infinity, and a secondary tube lens forms an image at the intermediate image plane. (This intermediate image is focused in turn by the ocular.) This configuration allows for a flexible distance between the objective and ocular because the distance between the objective and the tube lens can be varied without affecting the image-forming characteristics of the microscope. Note that objectives designed for infinity-corrected optics are not interchangeable with objectives designed for standard tube-length optics.

See Thin-film interference coating.

A type of illumination optics usually used in wide-field epifluorescence microscopes, in which the image of the light source is completely out of focus at the specimen plane.

An optical filter that attenuates shorter wavelengths and transmits longer wavelengths over the active range of the spectrum (which depends on the specific application). LP filters are denoted by the cut-on wavelength at 50% of peak transmission.

The region of the electromagnetic spectrum ranging in wavelength from approximately 8 to 14 microns. This range generally represents the atmospheric transmission window used for thermal detection and imaging of lower temperature sources.

See Radiance.

The region of the electromagnetic spectrum ranging in wavelength from approximately 3 to 8 microns. This range generally represents the atmospheric transmission window used for thermal detection and imaging of higher temperature sources, as well as molecular absorption detection and spectroscopy.

A unit of length commonly used for measuring the wavelength of light: 1 nm = 10 angstroms (A) = 10^-9 meters; and 1000 nm = 1 micron (μ) = 10^-6 meters.

The region of the electromagnetic spectrum ranging in wavelength from approximately 750 to 2500 nanometers.

An optical filter that attenuates light by an amount independent of the wavelength within the useful range of the filter. Metal-coated filters typically have a wider neutral range than glass filters and are more heat-tolerant.

An angle of incidence of zero degrees.

In the microscope, a measure of the effective cone-angle of light focused onto the specimen (NA of the condenser) or light captured by the objective (NA of the objective). A high value of NA improves both the brightness and the resolution of the image. (NA = N sin(θ), where N is the refractive index of the medium surrounding the specimen and θ is the half-cone angle of the light.)

A logarithmic unit of transmission: OD = -log (T), where T is the transmission (0 < T ≤ 1).  In terms of %T, OD = -log (%T/100).

See Polarization.

See Wedge.

When light is represented as a sinusoidal vibration of the electric field, the fraction of a full wave cycle at a particular point in space and time, defined in units of degrees or radians. Usually the point of interest is at the surface of a substrate or a thin-film coating, where an instantaneous phase shift can occur upon transmission or reflection of the incident wave.

The difference in phase shift between the S-plane and P-plane components of light when reflected from, or transmitted through, the surface of a substrate or a thin-film interference coating. The phase difference generally varies with the wavelength of the incident light. At wavelengths where there is zero phase difference, the surface or coating will not alter the polarization state of the incident light.

The instantaneous change in the phase of the incident light when light is reflected from, or transmitted through,  the surface of a substrate or a thin-film coating. For a dielectric material such as glass, the phase change of the reflected light will be either 0 or 180 degrees, depending on three factors: 1) the refractive indices of the incident and emergent media, 2) the angle of incidence, and 3) the polarization state of the incident light. For a thin-film interference coating, the value generally will be other than 0 or 180 degrees, and generally will vary with the wavelength of the incident light.

Also Fading. A photochemical reaction that causes the intensity of fluorescence to decrease with time.

Small breaks in the coating of an interference filter, usually caused by the presence of dust particles on the substrate during coating.

The orientation of the electric field vibrations of propagating light. These vibrations are transverse to the direction of propagation of the light and can be oriented at some angle around the axis of propagation. When the orientation of the vibrations changes rapidly and randomly in time, the light is said to be unpolarized. When the vibrations are restricted to one particular orientation angle over an extended length of time, the light is said to be plane-polarized. (Light can be partially as well as totally plane-polarized.) Elliptical polarization is a special case in which the orientation rotates around the direction of propagation over an extended length of time, at the same frequency as the propagation light. (If the amplitude of the vibrations are equal for all orientations, it is said to be circularly polarized.)

When light strikes a specular surface at non-normal incidence, the component of the electric field vibrations parallel to the plane of incidence of the surface (P-plane) generally behaves differently than the component perpendicular to the plane of incidence (S-plane). This causes a polarizing effect that is aligned orthogonally to the orientation of the surface.

A name for a dichroic beamsplitter that has multiple reflection bands and transmission regions.

In fluorescence microscopy, fluorescent emissions from the specimen itself rather than emissions from any fluorochromes present in the specimen. See also Autofluorescence.

A measure of how efficiently a fluorochrome converts absorbed radiation into emitted fluorescence.

Any chemical process that reduces the quantum efficiency of a fluorochrome in situ.

A measure of the radiometric brightness of a light source. Technically, the radiance is the radiant flux emitted per unit solid angle per unit area of the light source. A common unit is watts per steradian per square centimeter. Luminance is a measure of the brightness of a light source as perceived by the human eye (i.e., a photometric measure), commonly measured in candelas per square centimeter.

A measure of the distortion a plane wave of light experiences when reflected off of a surface, measured in fractions or multiples of a wavelength of visible light (usually 546 or 633 nm). In general the RWD will be different than the surface flatness for non-normal angles of incidence. For optical components with both transmitted and reflected light paths, such as cube  beamsplitters,  the RWD represents the total wavefront distortion of the reflected light path through the component.

A shift in the apparent position of the specimen occurs when an optical element is inserted or removed, adjusted, or switched with another element. This shift can occur in X/Y and/or Z/focus planes, depending on the nature and specifications of the optical element.

See Polarization.

A set of specifications for defining the maximum allowable size and number of scratches and digs on an optical surface. The scratch/dig values (e.g., 80/50) specify the scratch width in microns and the dig diameter in tens of microns, respectively. Although extensive evaluation procedures exist if rigorous standards must be maintained (military specification MIL-F-48616, for example), a qualitative visual assessment of the scratches and digs usually suffices.

See Indirect fluorescence.

An optical filter that attenuates longer wavelengths and transmits shorter wavelengths over the active range of the spectrum (which depends on the specific application). SP filters are denoted by the cut-off wavelength at 50% of peak transmission.

The region of the electromagnetic spectrum ranging in wavelength from approximately 1.4 to 3 microns, representing the longer-wavelength band of the NIR. This range includes the primary optical telecommunications bands. With the appropriate photodetector, imaging and analysis based on the unique infrared absorption spectra of molecules can be achieved using standard optical systems.

A measure of the brightness of the desired fluorescence (signal) relative to the brightness of the background (noise).

A measure of the sharpness of the transition from the transmitting region to the blocking region of a color filter.

An instrument used for measuring the excitation and emission spectra of a fluorescent substance.

An optical configuration employed by most microscopes in which the objective forms an image at the intermediate image plane. This intermediate image is focused in turn by the ocular. The distance between the nosepiece that holds the objective and the barrel that holds the ocular is fixed at a standard length of 160 mm so that objectives can be interchangeable between microscopes. See also Infinity-corrected.

The shift in wavelength between the peak excitation intensity and peak emission intensity of a fluorochrome.

The ground and polished piece of optical glass that is used as a base for the beamsplitter coating.

A measure of the deviation of the surface of an optical element from a perfect plane, usually measured in fractions or multiples of a wavelength of visible light (usually 546 or 633 nm).

Another term for S-plane polarization (short for “transverse-electric” mode). See Polarization.

A type of optical coating composed of a stack of microscopically thin layers of material. Although each material is intrinsically colorless, the reflections created at each interface between the layers combine through wave interference to selectively reflect some wavelengths of light and transmit others. Thin-film interference coatings are the main component of interference filters, which consist of one or more coatings separated by glass substrates and frequently one or more layers of filter glass.

Another term for P-plane polarization (short for “transverse-magnetic” mode). See Polarization.

A measure of the distortion a plane wave of light undergoes when transmitted through an optical element, measured in fractions or multiples of a wavelength of visible light (usually 546 or 633 nm).

The region of the electromagnetic spectrum ranges in wavelength from approximately 100 to 400 nanometers. Three distinct bands are: 1) near-UV, from 320 to 400 nm; 2) mid-UV, from 190 to 320 nm; and 3) vacuum-UV (VUV), below 190 nm. The terms UV-A and UV-B denote bands with distinct physiological effects: 320 to 380 nm and 280 to 320 nm, respectively.

Also Parallelism. A measure of the deviation of the outer surfaces of an optical element from perfect parallelism, usually measured in arc-minutes or arcseconds of angle.

An epifluorescence microscope in which the full field of view is illuminated, in contrast to a confocal epifluorescence microscope. The term brightfield is also used, but this might be confused with traditional diascopic brightfield illumination.