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![Scientists are working in the lab](https://upload.wikimedia.org/wikipedia/commons/8/8c/Scientists_are_working_in_the_lab.9.jpg)
The optical microscope, also referred to as a light microscope, is a type of
microscope
A 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 invisi ...
that commonly uses
visible light
Light or visible light is electromagnetic radiation that can be perceived by the human eye. Visible light is usually defined as having wavelengths in the range of 400–700 nanometres (nm), corresponding to frequencies of 750–420 te ...
and a system of
lenses
A lens is a transmissive optical device which focuses or disperses a light beam by means of refraction. A simple lens consists of a single piece of transparent material, while a compound lens consists of several simple lenses (''elements''), ...
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
Resolution(s) may refer to:
Common meanings
* Resolution (debate), the statement which is debated in policy debate
* Resolution (law), a written motion adopted by a deliberative body
* New Year's resolution, a commitment that an individual mak ...
and sample
contrast.
The object is placed on a stage and may be directly viewed through one or two
eyepiece
An eyepiece, or ocular lens, is a type of lens that is attached to a variety of optical devices such as telescopes and microscopes. It is named because it is usually the lens that is closest to the eye when someone looks through the device. The ...
s on the microscope. In high-power microscopes, both eyepieces typically show the same image, but with a
stereo microscope, slightly different images are used to create a 3-D effect. A camera is typically used to capture the image (
micrograph
A micrograph or photomicrograph is a photograph or digital image taken through a microscope or similar device to show a magnified image of an object. This is opposed to a macrograph or photomacrograph, an image which is also taken on a mic ...
).
The sample can be lit in a variety of ways. Transparent objects can be lit from below and solid objects can be lit with light coming through (
bright field) or around (
dark field
Dark-field microscopy (also called dark-ground microscopy) describes microscopy methods, in both light and electron microscopy, which exclude the unscattered beam from the image. As a result, the field around the specimen (i.e., where there is ...
) the objective lens.
Polarised light
Polarization ( also polarisation) is a property applying to transverse waves that specifies the geometrical orientation of the oscillations. In a transverse wave, the direction of the oscillation is perpendicular to the direction of motion of t ...
may be used to determine crystal orientation of metallic objects.
Phase-contrast imaging Phase-contrast imaging is a method of image, imaging that has a range of different applications. It exploits differences in the refractive index of different materials to differentiate between structures under analysis. In conventional Light microsc ...
can be used to increase image contrast by highlighting small details of differing refractive index.
A range of
objective
Objective may refer to:
* Objective (optics), an element in a camera or microscope
* ''The Objective'', a 2008 science fiction horror film
* Objective pronoun, a personal pronoun that is used as a grammatical object
* Objective Productions, a Brit ...
lenses with different magnification are usually provided mounted on a turret, allowing them to be rotated into place and providing an ability to zoom-in. The maximum magnification power of optical microscopes is typically limited to around 1000x because of the limited resolving power of visible light. While larger magnifications are possible no additional details of the object are resolved.
Alternatives to optical microscopy which do not use visible light include
scanning electron microscopy and
transmission electron microscopy
Transmission electron microscopy (TEM) is a microscopy technique in which a beam of electrons is transmitted through a specimen to form an image. The specimen is most often an ultrathin section less than 100 nm thick or a suspension on a g ...
and
scanning probe microscopy and as a result, can achieve much greater magnifications.
Types
![Microscope simple diagram](https://upload.wikimedia.org/wikipedia/commons/4/4f/Microscope_simple_diagram.png)
There are two basic types of optical microscopes: simple microscopes and compound microscopes. A simple microscope uses the
optical power
In optics, optical power (also referred to as dioptric power, refractive power, focusing power, or convergence power) is the degree to which a lens, mirror, or other optical system converges or diverges light. It is equal to the reciprocal of the ...
of single lens or group of lenses for magnification. A compound microscope uses a system of lenses (one set enlarging the image produced by another) to achieve much higher magnification of an object. The vast majority of modern
research
Research is "creativity, creative and systematic work undertaken to increase the stock of knowledge". It involves the collection, organization and analysis of evidence to increase understanding of a topic, characterized by a particular att ...
microscopes are compound microscopes while some cheaper commercial
digital microscopes are simple single lens microscopes. Compound microscopes can be further divided into a variety of other types of microscopes which differ in their optical configurations, cost, and intended purposes.
Simple microscope
A simple microscope uses a lens or set of lenses to enlarge an object through angular magnification alone, giving the viewer an erect enlarged
virtual image
In optics, an ''image'' is defined as the collection of focus points of light rays coming from an object. A '' real image'' is the collection of focus points made by converging rays, while a virtual image is the collection of focus points ma ...
. The use of a single convex lens or groups of lenses are found in simple magnification devices such as the
magnifying glass,
loupe
A loupe ( ) is a simple, small magnification device used to see small details more closely. They generally have higher magnification than a magnifying glass, and are designed to be held or worn close to the eye. A loupe does not have an attached h ...
s, and
eyepiece
An eyepiece, or ocular lens, is a type of lens that is attached to a variety of optical devices such as telescopes and microscopes. It is named because it is usually the lens that is closest to the eye when someone looks through the device. The ...
s for
telescope
A telescope is a device used to observe distant objects by their emission, absorption, or reflection of electromagnetic radiation. Originally meaning only an optical instrument using lenses, curved mirrors, or a combination of both to observ ...
s and microscopes.
Compound microscope
![Microscope compound diagram](https://upload.wikimedia.org/wikipedia/commons/f/fe/Microscope_compound_diagram.png)
A compound microscope uses a lens close to the object being viewed to collect light (called the
objective
Objective may refer to:
* Objective (optics), an element in a camera or microscope
* ''The Objective'', a 2008 science fiction horror film
* Objective pronoun, a personal pronoun that is used as a grammatical object
* Objective Productions, a Brit ...
lens) which focuses a
real image of the object inside the microscope (image 1). That image is then magnified by a second lens or group of lenses (called the
eyepiece
An eyepiece, or ocular lens, is a type of lens that is attached to a variety of optical devices such as telescopes and microscopes. It is named because it is usually the lens that is closest to the eye when someone looks through the device. The ...
) that gives the viewer an enlarged inverted virtual image of the object (image 2).
The use of a compound objective/eyepiece combination allows for much higher magnification. Common compound microscopes often feature exchangeable objective lenses, allowing the user to quickly adjust the magnification.
[ A compound microscope also enables more advanced illumination setups, such as ]phase contrast Phase-contrast imaging is a method of imaging that has a range of different applications. It exploits differences in the refractive index of different materials to differentiate between structures under analysis. In conventional light microscopy, ph ...
.
Other microscope variants
There are many variants of the compound optical microscope design for specialized purposes. Some of these are physical design differences allowing specialization for certain purposes:
* Stereo microscope, a low-powered microscope which provides a stereoscopic view of the sample, commonly used for dissection.
* Comparison microscope
A comparison microscope is a device used to analyze side-by-side specimens. It consists of two microscopes connected by an optical bridge, which results in a split view window enabling two separate objects to be viewed simultaneously. This avoids t ...
, which has two separate light paths allowing direct comparison of two samples via one image in each eye.
* Inverted microscope
An inverted microscope is a microscope with its light source and condenser on the top, above the stage pointing down, while the objectives and turret are below the stage pointing up. It was invented in 1850 by J. Lawrence Smith, a faculty membe ...
, for studying samples from below; useful for cell cultures in liquid, or for metallography.
* Fiber optic connector inspection microscope, designed for connector end-face inspection
* Traveling microscope
A travelling microscope is an instrument for measuring length with a resolution typically in the order of 0.01mm. The precision is such that better-quality instruments have measuring scales made from Invar to avoid misreadings due to thermal effe ...
, for studying samples of high optical resolution.
Other microscope variants are designed for different illumination techniques:
* Petrographic microscope
A petrographic microscope is a type of optical microscope used in petrology and optical mineralogy to identify rocks and minerals in thin sections. The microscope is used in optical mineralogy and petrography, a branch of petrology whi ...
, whose design usually includes a polarizing filter, rotating stage and gypsum plate to facilitate the study of minerals or other crystalline materials whose optical properties can vary with orientation.
* Polarizing microscope
A petrographic microscope is a type of optical microscope used in petrology and optical mineralogy to identify Rock (geology), rocks and minerals in thin sections. The microscope is used in optical mineralogy and petrography, a branch of ...
, similar to the petrographic microscope.
* Phase-contrast microscope
__NOTOC__
Phase-contrast microscopy (PCM) is an optical microscopy technique that converts phase shifts in light passing through a transparent specimen to brightness changes in the image. Phase shifts themselves are invisible, but become visible ...
, which applies the phase contrast illumination method.
* Epifluorescence microscope
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 microsc ...
, designed for analysis of samples which include fluorophores.
* Confocal microscope
Confocal microscopy, most frequently confocal laser scanning microscopy (CLSM) or laser confocal scanning microscopy (LCSM), is an optical imaging technique for increasing optical resolution and contrast of a micrograph by means of using a sp ...
, a widely used variant of epifluorescent illumination which uses a scanning laser to illuminate a sample for fluorescence.
* Two-photon microscope, used to image fluorescence deeper in scattering media and reduce photobleaching, especially in living samples.
* Student microscope – an often low-power microscope with simplified controls and sometimes low quality optics designed for school use or as a starter instrument for children.
* Ultramicroscope
An ultramicroscope is a microscope with a system that lights the object in a way that allows viewing of tiny particles via light scattering, and not light reflection or absorption. When the diameter of a particle is below or near the wavelength o ...
, an adapted light microscope that uses light scattering
Scattering is a term used in physics to describe a wide range of physical processes where moving particles or radiation of some form, such as light or sound, are forced to deviate from a straight trajectory by localized non-uniformities (including ...
to allow viewing of tiny particles whose diameter is below or near the wavelength of visible light (around 500 nanometers); mostly obsolete since the advent of electron microscope
An electron microscope is a microscope that uses a beam of accelerated electrons as a source of illumination. As the wavelength of an electron can be up to 100,000 times shorter than that of visible light photons, electron microscopes have a hi ...
s
* Tip-enhanced Raman microscope, is a variant of optical microscope based on tip-enhanced Raman spectroscopy Tip-enhanced Raman spectroscopy (TERS) is a variant of surface-enhanced Raman spectroscopy (SERS) that combines scanning probe microscopy with Raman spectroscopy. High spatial resolution chemical imaging is possible ''via'' TERS, with routine demons ...
, without traditional wavelength-based resolution limits. This microscope primarily realized on the scanning-probe microscope platforms using all optical tools.
Digital microscope
A digital microscope is a microscope equipped with a digital camera
A digital camera is a camera that captures photographs in digital memory. Most cameras produced today are digital, largely replacing those that capture images on photographic film. Digital cameras are now widely incorporated into mobile devices ...
allowing observation of a sample via a computer
A computer is a machine that can be programmed to Execution (computing), carry out sequences of arithmetic or logical operations (computation) automatically. Modern digital electronic computers can perform generic sets of operations known as C ...
. Microscopes can also be partly or wholly computer-controlled with various levels of automation. Digital microscopy allows greater analysis of a microscope image, for example, measurements of distances and areas and quantitation of a fluorescent or histological
Histology,
also known as microscopic anatomy or microanatomy, is the branch of biology which studies the microscopic anatomy of biological tissues. Histology is the microscopic counterpart to gross anatomy, which looks at larger structures vis ...
stain.
Low-powered digital microscopes, USB microscope
A USB microscope is a low-powered digital microscope which connects to a computer's USB port. Microscopes essentially the same as USB models are also available with other interfaces either in addition to or instead of USB, such as via WiFi ...
s, are also commercially available. These are essentially webcam
A webcam is a video camera which is designed to record or stream to a computer or computer network. They are primarily used in videotelephony, livestreaming and social media, and security. Webcams can be built-in computer hardware or peripheral ...
s with a high-powered macro lens
Macro photography (or photomacrography or macrography, and sometimes macrophotography) is extreme close-up photography, usually of very small subjects and living organisms like insects, in which the size of the subject in the photograph is grea ...
and generally do not use transillumination
Transillumination is the technique of sample illumination by transmission of light through the sample. Transillumination is used in a variety of methods of imaging.
Microscopy
In microscopy transillumination refers to the illumination of a sample ...
. The camera attached directly to the USB
Universal Serial Bus (USB) is an industry standard that establishes specifications for cables, connectors and protocols for connection, communication and power supply (interfacing) between computers, peripherals and other computers. A broad ...
port of a computer so that the images are shown directly on the monitor. They offer modest magnifications (up to about 200×) without the need to use eyepieces, and at very low cost. High power illumination is usually provided by an LED
A light-emitting diode (LED) is a semiconductor Electronics, device that Light#Light sources, emits light when Electric current, current flows through it. Electrons in the semiconductor recombine with electron holes, releasing energy i ...
source or sources adjacent to the camera lens.
Digital microscopy with very low light levels to avoid damage to vulnerable biological samples is available using sensitive photon-counting digital cameras. It has been demonstrated that a light source providing pairs of entangled photons may minimize the risk of damage to the most light-sensitive samples. In this application of ghost imaging
Ghost imaging (also called "coincidence imaging", "two-photon imaging" or "correlated-photon imaging") is a technique that produces an image of an object by combining information from two light detectors: a conventional, ''multi-pixel'' detecto ...
to photon-sparse microscopy, the sample is illuminated with infrared photons, each of which is spatially correlated with an entangled partner in the visible band for efficient imaging by a photon-counting camera.
History
Invention
The earliest microscopes were single lens
A lens is a transmissive optical device which focuses or disperses a light beam by means of refraction. A simple lens consists of a single piece of transparent material, while a compound lens consists of several simple lenses (''elements ...
magnifying glasses with limited magnification which date at least as far back as the widespread use of lenses in eyeglasses
Glasses, also known as eyeglasses or spectacles, are vision eyewear, with lenses (clear or tinted) mounted in a frame that holds them in front of a person's eyes, typically utilizing a bridge over the nose and hinged arms (known as temples or ...
in the 13th century.
Compound microscopes first appeared in Europe around 1620 including one demonstrated by Cornelis Drebbel in London (around 1621) and one exhibited in Rome in 1624.[Raymond J. Seeger, Men of Physics: Galileo Galilei, His Life and His Works, Elsevier - 2016, page 24][J. William Rosenthal, Spectacles and Other Vision Aids: A History and Guide to Collecting, Norman Publishing, 1996, page 391]
The actual inventor of the compound microscope is unknown although many claims have been made over the years. These include a claim 35 years after they appeared by Dutch
Dutch commonly refers to:
* Something of, from, or related to the Netherlands
* Dutch people ()
* Dutch language ()
Dutch may also refer to:
Places
* Dutch, West Virginia, a community in the United States
* Pennsylvania Dutch Country
People E ...
spectacle-maker Johannes Zachariassen that his father, Zacharias Janssen
Zacharias Janssen; also Zacharias Jansen or Sacharias Jansen; 1585 – pre-1632) was a Dutch spectacle-maker who lived most of his life in Middelburg. He is associated with the invention of the first optical telescope and/or the first truly ...
, invented the compound microscope and/or the telescope as early as 1590. Johannes' (some claim dubious) testimony pushes the invention date so far back that Zacharias would have been a child at the time, leading to speculation that, for Johannes' claim to be true, the compound microscope would have to have been invented by Johannes' grandfather, Hans Martens. Another claim is that Janssen's competitor, Hans Lippershey
Hans Lipperhey (circa 1570 – buried 29 September 1619), also known as Johann Lippershey or Lippershey, was a German- Dutch spectacle-maker. He is commonly associated with the invention of the telescope, because he was the first one who tried to ...
(who applied for the first telescope patent in 1608) also invented the compound microscope. Other historians point to the Dutch innovator Cornelis Drebbel with his 1621 compound microscope.
Galileo Galilei
Galileo di Vincenzo Bonaiuti de' Galilei (15 February 1564 – 8 January 1642) was an Italian astronomer, physicist and engineer, sometimes described as a polymath. Commonly referred to as Galileo, his name was pronounced (, ). He was ...
is also sometimes cited as a compound microscope inventor. After 1610, he found that he could close focus his telescope to view small objects, such as flies, close up and/or could look through the wrong end in reverse to magnify small objects. The only drawback was that his 2 foot long telescope had to be extended out to 6 feet to view objects that close. After seeing the compound microscope built by Drebbel exhibited in Rome in 1624, Galileo built his own improved version. In 1625, Giovanni Faber
Giovanni Faber (or Johann Faber, sometimes also known as Fabri or Fabro) (1574–1629) was a German papal doctor, botanist and art collector, originally from Bamberg in Bavaria, who lived in Rome from 1598. He was curator of the Vatican botan ...
coined the name ''microscope'' for the compound microscope Galileo submitted to the Accademia dei Lincei in 1624 (Galileo had called it the "''occhiolino''" or "''little eye''"). Faber coined the name from the Greek
Greek may refer to:
Greece
Anything of, from, or related to Greece, a country in Southern Europe:
*Greeks, an ethnic group.
*Greek language, a branch of the Indo-European language family.
**Proto-Greek language, the assumed last common ancestor ...
words ''μικρόν'' (micron) meaning "small", and ''σκοπεῖν'' (skopein) meaning "to look at", a name meant to be analogous with "telescope", another word coined by the Linceans.
Christiaan Huygens, another Dutchman, developed a simple 2-lens ocular system in the late 17th century that was achromatically corrected, and therefore a huge step forward in microscope development. The Huygens ocular is still being produced to this day, but suffers from a small field size, and other minor disadvantages.
Popularization
Antonie van Leeuwenhoek (1632–1724) is credited with bringing the microscope to the attention of biologists, even though simple magnifying lenses were already being produced in the 16th century. Van Leeuwenhoek's home-made microscopes were simple microscopes, with a single very small, yet strong lens. They were awkward in use, but enabled van Leeuwenhoek to see detailed images. It took about 150 years of optical development before the compound microscope was able to provide the same quality image as van Leeuwenhoek's simple microscopes, due to difficulties in configuring multiple lenses. In the 1850s, John Leonard Riddell
John Leonard Riddell (February 20, 1807 – October 7, 1865) was a science lecturer, botanist, geologist, medical doctor, chemist, microscopist, numismatist, politician, and science fiction author in the United States. He was born in Leyden, Massa ...
, Professor of Chemistry at Tulane University
Tulane University, officially the Tulane University of Louisiana, is a private university, private research university in New Orleans, Louisiana. Founded as the Medical College of Louisiana in 1834 by seven young medical doctors, it turned into ...
, invented the first practical binocular microscope while carrying out one of the earliest and most extensive American microscopic investigations of cholera
Cholera is an infection of the small intestine by some strains of the bacterium ''Vibrio cholerae''. Symptoms may range from none, to mild, to severe. The classic symptom is large amounts of watery diarrhea that lasts a few days. Vomiting and ...
.
Lighting techniques
While basic microscope technology and optics have been available for over 400 years it is much more recently that techniques in sample illumination were developed to generate the high quality images seen today.
In August 1893, August Köhler
August Karl Johann Valentin Köhler (4 March 1866 – 12 March 1948) was a German professor and early staff member of Carl Zeiss AG in Jena, Germany. He is best known for his development of the microscopy technique of Köhler illumination, an imp ...
developed Köhler illumination Köhler illumination is a method of specimen illumination used for transmitted and reflected light (trans- and epi-illuminated) optical microscopy. Köhler illumination acts to generate an even illumination of the sample and ensures that an image o ...
. This method of sample illumination gives rise to extremely even lighting and overcomes many limitations of older techniques of sample illumination. Before development of Köhler illumination the image of the light source, for example a lightbulb
An electric light, lamp, or light bulb is an electrical component that produces light. It is the most common form of artificial lighting. Lamps usually have a base made of ceramic, metal, glass, or plastic, which secures the lamp in the so ...
filament, was always visible in the image of the sample.
The Nobel Prize
The Nobel Prizes ( ; sv, Nobelpriset ; no, Nobelprisen ) are five separate prizes that, according to Alfred Nobel's will of 1895, are awarded to "those who, during the preceding year, have conferred the greatest benefit to humankind." Alfr ...
in physics was awarded to Dutch physicist Frits Zernike
Frits Zernike (; 16 July 1888 – 10 March 1966) was a Dutch physicist and winner of the Nobel Prize in Physics in 1953 for his invention of the phase-contrast microscope.
Early life and education
Frits Zernike was born on 16 July 1888 in Am ...
in 1953 for his development of phase contrast Phase-contrast imaging is a method of imaging that has a range of different applications. It exploits differences in the refractive index of different materials to differentiate between structures under analysis. In conventional light microscopy, ph ...
illumination which allows imaging of transparent samples. By using interference
Interference is the act of interfering, invading, or poaching. Interference may also refer to:
Communications
* Interference (communication), anything which alters, modifies, or disrupts a message
* Adjacent-channel interference, caused by extr ...
rather than absorption of light, extremely transparent samples, such as live mammalian
Mammals () are a group of vertebrate animals constituting the class Mammalia (), characterized by the presence of mammary glands which in females produce milk for feeding (nursing) their young, a neocortex (a region of the brain), fur o ...
cells, can be imaged without having to use staining techniques. Just two years later, in 1955, Georges Nomarski Georges (Jerzy) Nomarski (January 6, 1919 – 1997) was a Polish physicist and optics theoretician. Creator of differential interference contrast (DIC) microscopy, the method is widely used to study live biological specimens and unstained tissues a ...
published the theory for differential interference contrast
Differential interference contrast (DIC) microscopy, also known as Nomarski interference contrast (NIC) or Nomarski microscopy, is an optical microscopy technique used to enhance the contrast in unstained, transparent samples. DIC works on the p ...
microscopy, another interference
Interference is the act of interfering, invading, or poaching. Interference may also refer to:
Communications
* Interference (communication), anything which alters, modifies, or disrupts a message
* Adjacent-channel interference, caused by extr ...
-based imaging technique.
Fluorescence microscopy
Modern biological microscopy depends heavily on the development of fluorescent
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, ...
probes for specific structures within a cell. In contrast to normal transilluminated light microscopy, in fluorescence microscopy
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 microscop ...
the sample is illuminated through the objective lens with a narrow set of wavelengths of light. This light interacts with fluorophores in the sample which then emit light of a longer wavelength
In physics, the wavelength is the spatial period of a periodic wave—the distance over which the wave's shape repeats.
It is the distance between consecutive corresponding points of the same phase on the wave, such as two adjacent crests, tro ...
. It is this emitted light which makes up the image.
Since the mid-20th century chemical fluorescent stains, such as DAPI
DAPI (pronounced 'DAPPY', /ˈdæpiː/), 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 inta ...
which binds to DNA, have been used to label specific structures within the cell. More recent developments include immunofluorescence, which uses fluorescently labelled antibodies to recognise specific proteins within a sample, and fluorescent proteins like GFP GFP may refer to:
Organisations
* Gaelic Football Provence, a French Gaelic Athletic Association club
* Geheime Feldpolizei, the German secret military police during the Second World War
* French Group for the Study of Polymers and their Applicat ...
which a live cell can express making it fluorescent.
Components
All modern optical microscopes designed for viewing samples by transmitted light share the same basic components of the light path. In addition, the vast majority of microscopes have the same 'structural' components (numbered below according to the image on the right):
* Eyepiece (ocular lens) (1)
* Objective turret, revolver, or revolving nose piece (to hold multiple objective lenses) (2)
* Objective lenses
In optical engineering, the objective is the optical element that gathers light from the object being observed and focuses the light rays to produce a real image. Objectives can be a single lens or mirror, or combinations of several optical elem ...
(3)
* Focus knobs (to move the stage)
** Coarse adjustment (4)
** Fine adjustment (5)
* Stage (to hold the specimen) (6)
* Light source (a light
Light or visible light is electromagnetic radiation that can be perceived by the human eye. Visible light is usually defined as having wavelengths in the range of 400–700 nanometres (nm), corresponding to frequencies of 750–420 te ...
or a mirror
A mirror or looking glass is an object that reflects an image. Light that bounces off a mirror will show an image of whatever is in front of it, when focused through the lens of the eye or a camera. Mirrors reverse the direction of the im ...
) (7)
* Diaphragm and condenser (8)
* Mechanical stage (9)
Eyepiece (ocular lens)
The eyepiece
An eyepiece, or ocular lens, is a type of lens that is attached to a variety of optical devices such as telescopes and microscopes. It is named because it is usually the lens that is closest to the eye when someone looks through the device. The ...
, or ocular lens, is a cylinder containing two or more lenses; its function is to bring the image into focus for the eye. The eyepiece is inserted into the top end of the body tube. Eyepieces are interchangeable and many different eyepieces can be inserted with different degrees of magnification. Typical magnification values for eyepieces include 5×, 10× (the most common), 15× and 20×. In some high performance microscopes, the optical configuration of the objective lens and eyepiece are matched to give the best possible optical performance. This occurs most commonly with apochromat
An apochromat, or apochromatic lens (apo), is a photographic or other lens that has better correction of chromatic and spherical aberration than the much more common achromat lenses.
Explanation
Chromatic aberration is the phenomenon of differen ...
ic objectives.
Objective turret (revolver or revolving nose piece)
Objective turret, revolver, or revolving nose piece is the part that holds the set of objective lenses. It allows the user to switch between objective lenses.
Objective lens
At the lower end of a typical compound optical microscope, there are one or more objective lens
In optical engineering, the objective is the optical element that gathers light from the object being observed and focuses the light rays to produce a real image. Objectives can be a single lens or mirror, or combinations of several optical elem ...
es that collect light from the sample. The objective is usually in a cylinder housing containing a glass single or multi-element compound lens. Typically there will be around three objective lenses screwed into a circular nose piece which may be rotated to select the required objective lens. These arrangements are designed to be parfocal, which means that when one changes from one lens to another on a microscope, the sample stays in focus
Focus, or its plural form foci may refer to:
Arts
* Focus or Focus Festival, former name of the Adelaide Fringe arts festival in South Australia Film
*''Focus'', a 1962 TV film starring James Whitmore
* ''Focus'' (2001 film), a 2001 film based ...
. Microscope objectives are characterized by two parameters, namely, magnification and numerical aperture. The former typically ranges from 5× to 100× while the latter ranges from 0.14 to 0.7, corresponding to focal lengths of about 40 to 2 mm, respectively. Objective lenses with higher magnifications normally have a higher numerical aperture and a shorter depth of field in the resulting image. Some high performance objective lenses may require matched eyepieces to deliver the best optical performance.
Oil immersion objective
Some microscopes make use of oil-immersion objectives or water-immersion objectives for greater resolution at high magnification. These are used with index-matching material such as immersion oil
In light microscopy, oil immersion is a technique used to increase the resolving power of a microscope. This is achieved by immersing both the objective lens and the specimen in a transparent oil of high refractive index, thereby increasing the ...
or water and a matched cover slip between the objective lens and the sample. The refractive index of the index-matching material is higher than air allowing the objective lens to have a larger numerical aperture (greater than 1) so that the light is transmitted from the specimen to the outer face of the objective lens with minimal refraction. Numerical apertures as high as 1.6 can be achieved. The larger numerical aperture allows collection of more light making detailed observation of smaller details possible. An oil immersion lens usually has a magnification of 40 to 100×.
Focus knobs
Adjustment knobs move the stage up and down with separate adjustment for coarse and fine focusing. The same controls enable the microscope to adjust to specimens of different thickness. In older designs of microscopes, the focus adjustment wheels move the microscope tube up or down relative to the stand and had a fixed stage.
Frame
The whole of the optical assembly is traditionally attached to a rigid arm, which in turn is attached to a robust U-shaped foot to provide the necessary rigidity. The arm angle may be adjustable to allow the viewing angle to be adjusted.
The frame provides a mounting point for various microscope controls. Normally this will include controls for focusing, typically a large knurled wheel to adjust coarse focus, together with a smaller knurled wheel to control fine focus. Other features may be lamp controls and/or controls for adjusting the condenser.
Stage
The stage is a platform below the objective lens which supports the specimen being viewed. In the center of the stage is a hole through which light passes to illuminate the specimen. The stage usually has arms to hold slides (rectangular glass plates with typical dimensions of 25×75 mm, on which the specimen is mounted).
At magnifications higher than 100× moving a slide by hand is not practical. A mechanical stage, typical of medium and higher priced microscopes, allows tiny movements of the slide via control knobs that reposition the sample/slide as desired. If a microscope did not originally have a mechanical stage it may be possible to add one.
All stages move up and down for focus. With a mechanical stage slides move on two horizontal axes for positioning the specimen to examine specimen details.
Focusing starts at lower magnification in order to center the specimen by the user on the stage. Moving to a higher magnification requires the stage to be moved higher vertically for re-focus at the higher magnification and may also require slight horizontal specimen position adjustment. Horizontal specimen position adjustments are the reason for having a mechanical stage.
Due to the difficulty in preparing specimens and mounting them on slides, for children it is best to begin with prepared slides that are centered and focus easily regardless of the focus level used.
Light source
Many sources of light can be used. At its simplest, daylight is directed via a mirror
A mirror or looking glass is an object that reflects an image. Light that bounces off a mirror will show an image of whatever is in front of it, when focused through the lens of the eye or a camera. Mirrors reverse the direction of the im ...
. Most microscopes, however, have their own adjustable and controllable light source – often a halogen lamp
A halogen lamp (also called tungsten halogen, quartz-halogen, and quartz iodine lamp) is an incandescent lamp consisting of a tungsten filament sealed in a compact transparent envelope that is filled with a mixture of an inert gas and a small ...
, although illumination using LED
A light-emitting diode (LED) is a semiconductor Electronics, device that Light#Light sources, emits light when Electric current, current flows through it. Electrons in the semiconductor recombine with electron holes, releasing energy i ...
s and laser
A laser is a device that emits light through a process of optical amplification based on the stimulated emission of electromagnetic radiation. The word "laser" is an acronym for "light amplification by stimulated emission of radiation". The fir ...
s are becoming a more common provision. Köhler illumination Köhler illumination is a method of specimen illumination used for transmitted and reflected light (trans- and epi-illuminated) optical microscopy. Köhler illumination acts to generate an even illumination of the sample and ensures that an image o ...
is often provided on more expensive instruments.
Condenser
The condenser is a lens designed to focus light from the illumination source onto the sample. The condenser may also include other features, such as a diaphragm and/or filters, to manage the quality and intensity of the illumination. For illumination techniques like dark field
Dark-field microscopy (also called dark-ground microscopy) describes microscopy methods, in both light and electron microscopy, which exclude the unscattered beam from the image. As a result, the field around the specimen (i.e., where there is ...
, phase contrast Phase-contrast imaging is a method of imaging that has a range of different applications. It exploits differences in the refractive index of different materials to differentiate between structures under analysis. In conventional light microscopy, ph ...
and differential interference contrast
Differential interference contrast (DIC) microscopy, also known as Nomarski interference contrast (NIC) or Nomarski microscopy, is an optical microscopy technique used to enhance the contrast in unstained, transparent samples. DIC works on the p ...
microscopy additional optical components must be precisely aligned in the light path.
Magnification
The actual power or magnification of a compound optical microscope is the product of the powers of the eyepiece
An eyepiece, or ocular lens, is a type of lens that is attached to a variety of optical devices such as telescopes and microscopes. It is named because it is usually the lens that is closest to the eye when someone looks through the device. The ...
and the objective lens. For example a 10x eyepiece magnification and a 100x objective lens magnification gives a total magnification of 1,000×. Modified environments such as the use of oil or ultraviolet light can increase the resolution and allow for resolved details at magnifications larger than 1,000x.
Operation
Illumination techniques
Many techniques are available which modify the light path to generate an improved contrast image from a sample. Major techniques for generating increased contrast from the sample include cross-polarized light, dark field
Dark-field microscopy (also called dark-ground microscopy) describes microscopy methods, in both light and electron microscopy, which exclude the unscattered beam from the image. As a result, the field around the specimen (i.e., where there is ...
, phase contrast Phase-contrast imaging is a method of imaging that has a range of different applications. It exploits differences in the refractive index of different materials to differentiate between structures under analysis. In conventional light microscopy, ph ...
and differential interference contrast
Differential interference contrast (DIC) microscopy, also known as Nomarski interference contrast (NIC) or Nomarski microscopy, is an optical microscopy technique used to enhance the contrast in unstained, transparent samples. DIC works on the p ...
illumination. A recent technique (Sarfus
Sarfus is an optical quantitative imaging technique based on the association of:
*an upright or inverted optical microscope in crossed polarization configuration and
*specific supporting plates – called surfs – on which the sample to observe i ...
) combines cross-polarized light and specific contrast-enhanced slides for the visualization of nanometric samples.
tissue paper
Tissue paper or simply tissue is a lightweight paper or, light crêpe paper. Tissue can be made from recycled paper pulp on a paper machine.
Tissue paper is very versatile, and different kinds of tissue are made to best serve these purposes, w ...
. 1.559 μm/pixel." align="center">
File:Paper Micrograph Bright.png,
Bright field illumination, sample contrast comes from
absorbance
Absorbance is defined as "the logarithm of the ratio of incident to transmitted radiant power through a sample (excluding the effects on cell walls)". Alternatively, for samples which scatter light, absorbance may be defined as "the negative lo ...
of light in the sample.
File:Paper Micrograph Cross-Polarised.png,
Cross-polarized light illumination, sample contrast comes from rotation of
polarized light through the sample.
File:Paper Micrograph Dark.png,
Dark field
Dark-field microscopy (also called dark-ground microscopy) describes microscopy methods, in both light and electron microscopy, which exclude the unscattered beam from the image. As a result, the field around the specimen (i.e., where there is ...
illumination, sample contrast comes from light
scattered by the sample.
File:Paper Micrograph Phase.png,
Phase contrast Phase-contrast imaging is a method of imaging that has a range of different applications. It exploits differences in the refractive index of different materials to differentiate between structures under analysis. In conventional light microscopy, ph ...
illumination, sample contrast comes from
interference
Interference is the act of interfering, invading, or poaching. Interference may also refer to:
Communications
* Interference (communication), anything which alters, modifies, or disrupts a message
* Adjacent-channel interference, caused by extr ...
of different path lengths of light through the sample.
Other techniques
Modern microscopes allow more than just observation of transmitted light image of a sample; there are many techniques which can be used to extract other kinds of data. Most of these require additional equipment in addition to a basic compound microscope.
* Reflected light, or incident, illumination (for analysis of surface structures)
* Fluorescence microscopy, both:
:*
Epifluorescence microscopy
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 microsc ...
:*
Confocal microscopy
Confocal microscopy, most frequently confocal laser scanning microscopy (CLSM) or laser confocal scanning microscopy (LCSM), is an optical imaging technique for increasing optical resolution and contrast of a micrograph by means of using a s ...
*
Microspectroscopy (where a UV-visible spectrophotometer is integrated with an optical microscope)
* Ultraviolet microscopy
* Near-Infrared microscopy
* Multiple transmission microscopy for contrast enhancement and aberration reduction.
* Automation (for automatic scanning of a large sample or image capture)
Applications
![Vaginal wet mount of candidal vulvovaginitis](https://upload.wikimedia.org/wikipedia/commons/3/38/Vaginal_wet_mount_of_candidal_vulvovaginitis.jpg)
Optical microscopy is used extensively in microelectronics, nanophysics, biotechnology, pharmaceutic research, mineralogy and microbiology.
Optical microscopy is used for
medical diagnosis
Medical diagnosis (abbreviated Dx, Dx, or Ds) is the process of determining which disease or condition explains a person's symptoms and signs. It is most often referred to as diagnosis with the medical context being implicit. The information r ...
, the field being termed
histopathology when dealing with tissues, or in
smear test
Cytopathology (from Greek , ''kytos'', "a hollow"; , ''pathos'', "fate, harm"; and , ''-logia'') is a branch of pathology that studies and diagnoses diseases on the cellular level. The discipline was founded by George Nicolas Papanicolaou in 1 ...
s on free cells or tissue fragments.
In industrial use, binocular microscopes are common. Aside from applications needing true
depth perception
Depth perception is the ability to perceive distance to objects in the world using the visual system and visual perception. It is a major factor in perceiving the world in three dimensions. Depth perception happens primarily due to stereopsis an ...
, the use of dual eyepieces reduces
eye strain associated with long workdays at a microscopy station. In certain applications, long-working-distance or long-focus microscopes
are beneficial. An item may need to be examined behind a
window
A window is an opening in a wall, door, roof, or vehicle that allows the exchange of light and may also allow the passage of sound and sometimes air. Modern windows are usually glazed or covered in some other transparent or translucent mat ...
, or industrial subjects may be a hazard to the objective. Such optics resemble telescopes with close-focus capabilities.
Measuring microscopes are used for precision measurement. There are two basic types.
One has a
reticle
A reticle, or reticule also known as a graticule, is a pattern of fine lines or markings built into the eyepiece of an optical device such as a telescopic sight, spotting scope, theodolite, optical microscope or the screen of an oscilloscop ...
graduated to allow measuring distances in the focal plane. The other (and older) type has simple
crosshairs
A reticle, or reticule also known as a graticule, is a pattern of fine lines or markings built into the eyepiece of an optical device such as a telescopic sight, spotting scope, theodolite, optical microscope or the screen of an oscilloscope ...
and a micrometer mechanism for moving the subject relative to the microscope.
Very small, portable microscopes have found some usage in places where a laboratory microscope would be a burden.
Limitations
![Ernst-Abbe-Denkmal Jena Fürstengraben - 20140802 125709](https://upload.wikimedia.org/wikipedia/commons/4/46/Ernst-Abbe-Denkmal_Jena_F%C3%BCrstengraben_-_20140802_125709.jpg)
At very high magnifications with transmitted light, point objects are seen as fuzzy discs surrounded by
diffraction rings. These are called
Airy disk
In optics, the Airy disk (or Airy disc) and Airy pattern are descriptions of the best- focused spot of light that a perfect lens with a circular aperture can make, limited by the diffraction of light. The Airy disk is of importance in physics, ...
s. The ''resolving power'' of a microscope is taken as the ability to distinguish between two closely spaced Airy disks (or, in other words the ability of the microscope to reveal adjacent structural detail as distinct and separate). It is these impacts of diffraction that limit the ability to resolve fine details. The extent and magnitude of the diffraction patterns are affected by both the
wavelength
In physics, the wavelength is the spatial period of a periodic wave—the distance over which the wave's shape repeats.
It is the distance between consecutive corresponding points of the same phase on the wave, such as two adjacent crests, tro ...
of
light
Light or visible light is electromagnetic radiation that can be perceived by the human eye. Visible light is usually defined as having wavelengths in the range of 400–700 nanometres (nm), corresponding to frequencies of 750–420 te ...
(λ), the refractive materials used to manufacture the objective lens and the
numerical aperture (NA) of the objective lens. There is therefore a finite limit beyond which it is impossible to resolve separate points in the objective field, known as the
diffraction limit
The resolution of an optical imaging system a microscope, telescope, or camera can be limited by factors such as imperfections in the lenses or misalignment. However, there is a principal limit to the resolution of any optical system, due to t ...
. Assuming that optical aberrations in the whole optical set-up are negligible, the resolution ''d'', can be stated as:
:
Usually a wavelength of 550 nm is assumed, which corresponds to
green
Green is the color between cyan and yellow on the visible spectrum. It is evoked by light which has a dominant wavelength of roughly 495570 nm. In subtractive color systems, used in painting and color printing, it is created by a combi ...
light. With
air
The atmosphere of Earth is the layer of gases, known collectively as air, retained by Earth's gravity that surrounds the planet and forms its planetary atmosphere. The atmosphere of Earth protects life on Earth by creating pressure allowing f ...
as the external medium, the highest practical ''NA'' is 0.95, and with oil, up to 1.5. In practice the lowest value of ''d'' obtainable with conventional lenses is about 200 nm. A new type of lens using multiple scattering of light allowed to improve the resolution to below 100 nm.
Surpassing the resolution limit
Multiple techniques are available for reaching resolutions higher than the transmitted light limit described above. Holographic techniques, as described by Courjon and Bulabois in 1979, are also capable of breaking this resolution limit, although resolution was restricted in their experimental analysis.
Using fluorescent samples more techniques are available. Examples include
Vertico SMI,
near field scanning optical microscopy
Near-field scanning optical microscopy (NSOM) or scanning near-field optical microscopy (SNOM) is a microscopy technique for nanostructure investigation that breaks the far field Diffraction-limited system, resolution limit by exploiting the prop ...
which uses
evanescent waves
In electromagnetics, an evanescent field, or evanescent wave, is an oscillating electric and/or magnetic field that does not propagate as an electromagnetic wave but whose energy is spatially concentrated in the vicinity of the source (oscillati ...
, and
stimulated emission depletion. In 2005, a microscope capable of detecting a single molecule was described as a teaching tool.
Despite significant progress in the last decade, techniques for surpassing the diffraction limit remain limited and specialized.
While most techniques focus on increases in lateral resolution there are also some techniques which aim to allow analysis of extremely thin samples. For example,
sarfus
Sarfus is an optical quantitative imaging technique based on the association of:
*an upright or inverted optical microscope in crossed polarization configuration and
*specific supporting plates – called surfs – on which the sample to observe i ...
methods place the thin sample on a contrast-enhancing surface and thereby allows to directly visualize films as thin as 0.3 nanometers.
On 8 October 2014, the
Nobel Prize in Chemistry
)
, image = Nobel Prize.png
, alt = A golden medallion with an embossed image of a bearded man facing left in profile. To the left of the man is the text "ALFR•" then "NOBEL", and on the right, the text (smaller) "NAT•" then "M ...
was awarded to
Eric Betzig
Robert Eric Betzig (born January 13, 1960) is an American physicist who works as a professor of physics and professor of molecular and cell biology at the University of California, Berkeley. He is also a senior fellow at the Janelia Farm Research ...
,
William Moerner
William Esco Moerner (born June 24, 1953) is an American physical chemist and chemical physicist with current work in the biophysics and imaging of single molecules. He is credited with achieving the first optical detection and spectroscopy of a ...
and
Stefan Hell
Stefan Walter Hell HonFRMS (: born 23 December 1962) is a Romanian-German physicist and one of the directors of the Max Planck Institute for Biophysical Chemistry in Göttingen, Germany. He received the Nobel Prize in Chemistry in 2014 "for the d ...
for the development of super-resolved
fluorescence microscopy
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 microscop ...
.
Structured illumination SMI
SMI (spatially modulated illumination microscopy) is a light optical process of the so-called
point spread function (PSF) engineering. These are processes which modify the PSF of a
microscope
A 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 invisi ...
in a suitable manner to either increase the optical resolution, to maximize the precision of
distance
Distance is a numerical or occasionally qualitative measurement of how far apart objects or points are. In physics or everyday usage, distance may refer to a physical length or an estimation based on other criteria (e.g. "two counties over"). ...
measurements of fluorescent objects that are small relative to the
wavelength
In physics, the wavelength is the spatial period of a periodic wave—the distance over which the wave's shape repeats.
It is the distance between consecutive corresponding points of the same phase on the wave, such as two adjacent crests, tro ...
of the illuminating light, or to extract other structural parameters in the nanometer range.
Localization microscopy SPDMphymod
![3D Dual Color Super Resolution Microscopy Cremer 2010](https://upload.wikimedia.org/wikipedia/commons/1/1a/3D_Dual_Color_Super_Resolution_Microscopy_Cremer_2010.png)
SPDM (spectral precision distance microscopy), the basic localization microscopy technology is a light optical process of
fluorescence microscopy
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 microscop ...
which allows position, distance and angle measurements on "optically isolated" particles (e.g. molecules) well below the theoretical
limit of resolution for light microscopy. "Optically isolated" means that at a given point in time, only a single particle/molecule within a region of a size determined by conventional optical resolution (typically approx. 200–250 nm
diameter
In geometry, a diameter of a circle is any straight line segment that passes through the center of the circle and whose endpoints lie on the circle. It can also be defined as the longest chord of the circle. Both definitions are also valid fo ...
) is being registered. This is possible when
molecules within such a region all carry different spectral markers (e.g. different colors or other usable differences in the
light emission
This is a list of sources of light, the visible part of the electromagnetic spectrum. Light sources produce photons from another energy source, such as heat, chemical reactions, or conversion of mass or a different frequency of electromagnetic ener ...
of different particles).
Many standard fluorescent dyes like
GFP GFP may refer to:
Organisations
* Gaelic Football Provence, a French Gaelic Athletic Association club
* Geheime Feldpolizei, the German secret military police during the Second World War
* French Group for the Study of Polymers and their Applicat ...
, Alexa dyes, Atto dyes, Cy2/Cy3 and fluorescein molecules can be used for localization microscopy, provided certain photo-physical conditions are present. Using this so-called SPDMphymod (physically modifiable fluorophores) technology a single laser wavelength of suitable intensity is sufficient for nanoimaging.
3D super resolution microscopy
3D super resolution microscopy with standard fluorescent dyes can be achieved by combination of localization microscopy for standard fluorescent dyes SPDMphymod and structured illumination SMI.
STED
Stimulated emission depletion is a simple example of how higher resolution surpassing the diffraction limit is possible, but it has major limitations. STED is a fluorescence microscopy technique which uses a combination of light pulses to induce fluorescence in a small sub-population of fluorescent molecules in a sample. Each molecule produces a diffraction-limited spot of light in the image, and the centre of each of these spots corresponds to the location of the molecule. As the number of fluorescing molecules is low the spots of light are unlikely to overlap and therefore can be placed accurately. This process is then repeated many times to generate the image.
Stefan Hell
Stefan Walter Hell HonFRMS (: born 23 December 1962) is a Romanian-German physicist and one of the directors of the Max Planck Institute for Biophysical Chemistry in Göttingen, Germany. He received the Nobel Prize in Chemistry in 2014 "for the d ...
of the Max Planck Institute for Biophysical Chemistry was awarded the 10th German Future Prize in 2006 and Nobel Prize for Chemistry in 2014 for his development of the STED microscope and associated methodologies.
Alternatives
In order to overcome the limitations set by the diffraction limit of visible light other microscopes have been designed which use other waves.
*
Atomic force microscope
Atomic force microscopy (AFM) or scanning force microscopy (SFM) is a very-high-resolution type of scanning probe microscopy (SPM), with demonstrated resolution on the order of fractions of a nanometer, more than 1000 times better than the op ...
(AFM)
*
Scanning electron microscope (SEM)
*
Scanning ion-conductance microscopy
Scanning ion-conductance microscopy (SICM) is a scanning probe microscopy technique that uses an electrode as the probe tip. SICM allows for the determination of the surface topography of micrometer and even nanometer-range structures in aqueous m ...
(SICM)
*
Scanning tunneling microscope
A scanning tunneling microscope (STM) is a type of microscope used for imaging surfaces at the atomic level. Its development in 1981 earned its inventors, Gerd Binnig and Heinrich Rohrer, then at IBM Zürich, the Nobel Prize in Physics in 1986 ...
(STM)
*
Transmission electron microscopy
Transmission electron microscopy (TEM) is a microscopy technique in which a beam of electrons is transmitted through a specimen to form an image. The specimen is most often an ultrathin section less than 100 nm thick or a suspension on a g ...
(TEM)
* Ultraviolet microscope
*
X-ray microscope
An X-ray microscope uses electromagnetic radiation in the soft X-ray band to produce magnified images of objects. Since X-rays penetrate most objects, there is no need to specially prepare them for X-ray microscopy observations.
Unlike visible li ...
It is important to note that higher frequency waves have limited interaction with matter, for example soft tissues are relatively transparent to X-rays resulting in distinct sources of contrast and different target applications.
The use of electrons and X-rays in place of light allows much higher resolution – the wavelength of the radiation is shorter so the diffraction limit is lower. To make the
short-wavelength probe non-destructive, the atomic beam imaging system (
atomic nanoscope
The atomic de Broglie microscope (also atomic nanoscope, neutral beam microscope, or scanning helium microscope when helium is used as the probing atom) is an imaging system which is expected to provide resolution at the nanometer scale. It is som ...
) has been proposed and widely discussed in the literature, but it is not yet competitive with conventional imaging systems.
STM and AFM are scanning probe techniques using a small probe which is scanned over the sample surface. Resolution in these cases is limited by the size of the probe; micromachining techniques can produce probes with tip radii of 5–10 nm.
Additionally, methods such as electron or X-ray microscopy use a vacuum or partial vacuum, which limits their use for live and biological samples (with the exception of an
environmental scanning electron microscope
The environmental scanning electron microscope (ESEM) is a scanning electron microscope (SEM) that allows for the option of collecting electron micrographs of specimens that are wet, uncoated, or both by allowing for a gaseous environment in ...
). The specimen chambers needed for all such instruments also limits sample size, and sample manipulation is more difficult. Color cannot be seen in images made by these methods, so some information is lost. They are however, essential when investigating molecular or atomic effects, such as
age hardening
Precipitation hardening, also called age hardening or particle hardening, is a heat treatment technique used to increase the yield strength of malleable materials, including most structural alloys of aluminium, magnesium, nickel, titanium, and so ...
in
aluminium alloy
An aluminium alloy (or aluminum alloy; see spelling differences) is an alloy in which aluminium (Al) is the predominant metal. The typical alloying elements are copper, magnesium, manganese, silicon, tin, nickel and zinc. There are two principal ...
s, or the
microstructure
Microstructure is the very small scale structure of a material, defined as the structure of a prepared surface of material as revealed by an optical microscope above 25× magnification. The microstructure of a material (such as metals, polymers ...
of
polymers
A polymer (; Greek '' poly-'', "many" + ''-mer'', "part")
is a substance or material consisting of very large molecules called macromolecules, composed of many repeating subunits. Due to their broad spectrum of properties, both synthetic an ...
.
See also
*
Digital microscope
*
Köhler illumination Köhler illumination is a method of specimen illumination used for transmitted and reflected light (trans- and epi-illuminated) optical microscopy. Köhler illumination acts to generate an even illumination of the sample and ensures that an image o ...
*
Microscope slide
A microscope slide is a thin flat piece of glass, typically 75 by 26 mm (3 by 1 inches) and about 1 mm thick, used to hold objects for examination under a microscope. Typically the object is mounted (secured) on the slide, and then b ...
References
Cited sources
*
Further reading
* "Metallographic and Materialographic Specimen Preparation, Light Microscopy, Image Analysis and Hardness Testing", Kay Geels in collaboration with Struers A/S, ASTM International 2006.
"Light Microscopy: An ongoing contemporary revolution" Siegfried Weisenburger and Vahid Sandoghdar, arXiv:1412.3255 2014.
External links
Antique Microscopes & Scientific Instruments A site about Antique Microscopes, their Accessories, and History
Antique Microscopes.comA collection of early microscopes
an illustrated collection with more than 3000 photos of scientific microscopes by European makers
The Golub Collection A collection of 17th through 19th century microscopes, including extensive descriptions
concepts in optical microscopy
Online tutorial of practical optical microscopyat University of Cambridge
OpenWetWareCell Centered Database
{{DEFAULTSORT:Optical Microscope
Microscopes
Dutch inventions
Optical microscopy