Kinds of light microscopy

Depending on the properties of the object light changes its physical properties color (wavelength), brightness (the wave amplitude, phase, used in modern microscopes to create contrast.
For microscopy painted objects are the simplest known microscopes - the so-called normal (Fig. 1). As the eyes and the photographic plate easily capture differences in the color of the last beam, do not require any additional effort for consideration and shooting color image. The color image has a huge cognitive value for the interaction of the object with the paint can be judged on its chemical nature. Changing the paint mode, introducing additional processing solvents, clamps, coagulants and other substances (see Histological techniques), to achieve a very precise, including quantitative knowledge about the nature of the object and its chemistry. Based on this rapidly developing histo - and cytochemistry, which is unthinkable without microscopy. Many diagnostic techniques practical medicine is based on microscopy painted objects. In pathohistology, infectious diseases, Parasitology apply such specific dyes, methods of processing of specimens and modes of painting, which is often one look in the microscope for diagnosis or assessment of treatment results.
The most susceptible to staining fixed killed by drugs. Such fixed medications can be with high accuracy examined and photographed through a microscope, but they do not allow to evaluate the various forms of life microscopium object (move, merge, phagocytosis and other). Known dyes that are associated with living cells without disturbing their life.
Vital (lifetime) microscopy shows that many of the structure of the living cell relatively little change with skillful fixing and subsequent staining. This confirms the high scientific value of the information obtained by means of microscopy painted objects. Vital microscopy possible without staining, if a conventional microscope to introduce so-called dark-field condenser. He illuminates the object so that in the eye of the observer are only those rays that were scattered on the particles of the object and thereby changed the direction of its distribution. The rays that have passed through the background without scattering, eyes not fall. Therefore particles of the object Shine and stand out vividly against a dark background (dark field). Particles of the object are clearly visible, even if their size is less than the permitted range.
Dark-field microscopy provides the greatest possible contrast of the image, but the clarity of its useful increased considerably lower than conventional Meters dark-field Meters has been successfully applied to study spirohet, leptospirs, and other poorly painted microorganisms. When working with histological preparations it is not applicable.
Technically independent option dark-field microscopy is ultramicroscopy, in which the smallest studied particles highlights powerful side beam of light and visible points on a black background. Ultramicroscopy allows to count particles, to estimate their size and other properties. Apply for the study of colloidal solutions, aerosols, suspensions.
In recent years, dark-field microscopy was used less and less, because there are two new types of contrasting devices with much better performance - phase-contrast (Fig. 2, a and b) and the amplitude-contrast microscope. Technically they are similar, but they use different changes of the light beam in the object. The beam that has passed through the background of the sample, in the ideal case undergoes no changes. He goes through exactly certain parts of the lens. The beam that passes through an object undergoes diffraction, i.e. breaks up into bundles of diminishing intensity, which leave the object from different angles. Other properties of the beam (amplitude, wavelength, phase) change in varying degrees depending on the features of the object.
Almost all living microscopic objects look in the ordinary microscope barely visible, transparent, because they almost do not change either the amplitude or the color of the last beam through them.
They change only the phase of the wave, but this change was not revealed neither the eye nor a photographic plate. Beam, diffracted object and offset them by the phase that passes through those parts of the lens, where you can't go straight, nederevyannyy rays background. Practically it is not difficult to define, there where exactly will these rays. If you cover this area one of the lenses of the lens semi-transparent plate, capable of changing the phase, intensity, color or all three of these properties together, the background image will change its phase will decrease the brightness or will change color. The rays that have passed through the object, and rejected (dragerware) them, will pass under the lens of the record and, therefore, will not get those properties acquired through the plate, beams background. In the result of the difference between the rays of the background and the object will increase. If the phase difference between the rays of the background and the object reaches J/4 wavelength, in the final image is visible to the eye and plates contrast: dark object on a light background, or Vice versa, depending on the structure of the plate, which in this case is called "phase". If the record changes mainly the brightness and color of the background, such a microscope should be called amplitude-contrast (received widespread shorter, though not quite correct "anopthalmia"). Thus, the difference between the phase-contrast and amplitude-contrast microscope is determined by the properties of the plate in the lens that alters the properties nederevyannyy ray background. Image built these microscopes, much brighter and richer parts (Fig. 3 and 4)than dark-field painting.
With the advent of the phase and amplitude-contrast microscopes vital microscopy received an excellent technical and methodological base, the possibilities of which are close to the limit for the light optics. No commit or color of the object, these devices do not require. Modern vital M extremely expanded our knowledge about the behavior and dynamics of living micro-objects in natural and laboratory conditions and experiment. Rapid (rapid) and slow (tetraptera) microcinema made available for research of the processes, the flow rate too high or too low for visual observation.
Produced by industry phase-contrast and amplitude-contrast (anophthalmia) devices are inexpensive, easy to mount on serial microscopes; their use is no problem. These devices will no doubt find new applications both in research and in medical practice.


Ultraviolet microscopy is based on the ability of some substances selectively absorb ultraviolet light with the wavelength. This allows you to demonstrate and explore, including quantitatively, the distribution of substances in living cells or fixed preparations. For example, proteins and nucleic acids are equally transparent to visible light; considering an empty cell in visible light, it is impossible to determine where a protein or nucleic acid. But the ultraviolet rays of a certain length nucleic acid absorbs much more than protein. Therefore, in the ultraviolet microscope section containing nucleic acid, looks darker. Because ultraviolet rays directly not perceived by the eye, it is necessary to apply special converters of light. Ultraviolet microscopy technically much more difficult than light, its equipment is more expensive and methodology thinner. Despite this, its application is justified, as the scientific importance quick topographic description of the chemical composition of living cells is very high.
Much more affordable and promising fluorescence microscopy (see), which are widely used now in research and clinical diagnostic laboratories. While living object is treated with special dyes that are being lit in blue, purple or UV light begin to Shine, Shine with longer wave (green, yellow). Color excited secondary chemiluminescence depends on the chemical properties of an object and put him in the dye.
Polarizing microscopy based on the change of plane oscillations of a light wave after passing through the crystal. In practical medicine is not applied.
Modern Meters requires the use of a variety of auxiliary equipment. Heating tables and thermostats allow to stand and watch object for a long time at a given temperature. For long-term cultivation of microbes or tissue cultures in view of strong lens serve a variety of microcamera. The eyepiece and objective micrometers enable accurate measurement of micro-objects. Industry produces micromanipulators (see) for operations on the objects. To obtain a stereoscopic image at magnifications up to 100 times are binocular loupes (see) and stereo (Fig. 5). Widely produced and used equipment for microphotography and microinnochange (Fig. 6). Cm. also Microscopic techniques.