Light microscope how does it work

The optical or light microscope uses visible light transmitted through, refracted around, or reflected from a specimen. Light waves are chaotic; an incandescent light source emits light waves traveling in different paths and of varying wavelengths. Some of the lenses in a microscope bend these light waves into parallel paths, magnify and focus the light at the ocular. The power to enlarge the image of the specimen when viewed through a microscope is known as the magnification and is dependent upon how much the lenses bend the light waves.

Magnification is expressed in numeric multiples of how much enlargement occurs with a lens. If the magnification of a lens is 2X then it roughly doubles the size of the image of the object. With a compound microscope, the total magnification can be determined by multiplying the magnifications of the objective and ocular lenses. Consequently, an ocular lens of 10X coupled with a 40X objective yields a total magnification of X.

However, the higher the magnification the closer the lens must be to the specimen. Since a higher magnification lens bends light more severely, the specimen is brought into focus a shorter distance from the lens and this is known as the focal length.

Generally, a lens providing higher magnification will also provide better resolution. These two factors working together are very important in determining how a microscope works? The resolution of a specimen is highly dependent upon the light waves. The shortest distance between two points that the microscope can define as clearly being separate points is the resolution of the microscope.

Resolution is perhaps more important than magnification in understanding how a microscope works? If the points cannot be clearly focused then they are closer together than the resolution of the microscope and, regardless of the magnification, the image quality will be poor.

The resolution is determined by the frequency of the light waves illuminating the specimen and the quality of the lens. A rule of optical physics is that the shorter the wave length the greater the resolution. Usually expressed in microns, the best resolution a light microscope can produce is 0.

Discounting the light source, a lens having a resolution of 0. Contrast is another important ingredient in how a microscope works. If all of the light passes through a cell no details will be visible. Some light frequencies must be absorbed to different degrees by structures inside the cell and this allows you to see the specimen. A narrow beam provides higher contrast. Staining the specimen may be necessary to obtain the contrast you need to view the details of your sample.

As previously mentioned, optical microscopes are limited in resolution by the frequency of the light waves. Electron guns emit a flow of electrons of a considerably shorter wave length than visible light and this fact allows an electron microscope to have higher resolution and magnification.

In many ways, an electron microscope functions similarly to an optical scope except that, instead of visible light, a stream of electrons is used to illuminate the specimen. The electron beam is focused with magnetic lenses. Changes to the electron beam inside the specimen are recorded and an image is formed based upon these changes.

The objective lens magnifies the sample, as do the eyepieces you are looking through. Light Microscope Features and Functions. In order to focus the image, the coarse focusing is used first in order to put the sample in the correct location to obtain a clear image.

On light microscopes, moving the focusing knob will either move the stage up and down, or move the head of the microscope up and down. On the U2 biological microscope shown above, moving the coarse focus adjusts the height of the stage.

On most high school microscopes , the focusing mechanisms moves the height of the head of the microscope. Condenser Lens - collects the light from the illuminator and focuses it on the specimen.

Eyepiece Adjustment Knobs - used to bring the specimen into focus. Stage clips - used to keep the slide in place. Brightfield microscope. Best for Students. Most microscopes used in classrooms are bright field microscopes.

Bright field microscopy is the simplest form of optical microscopy illumination techniques. The term is derived from the fact that the specimen appears darker in contrast to the bright background.

Light from the illuminator is collected by the condenser and focused at the specimen mounted on the space. The light that passes through the specimen then goes through the objective lenses and ultimately through the eyepiece.

The specimen can either be stained or colorless. The pigmentation creates contrast which allows the viewer to see the image of the object being observed. This conventional technique is most suitable for observing the natural colors of the specimen. Phase-Contrast microscope. For the purpose of viewing structures e. This contrast-enhancing optical technique makes use of the minute differences in phase to create high-contrast images of an unstained specimen.

Phase-contrast microscopy employs special phase-contrast objectives and condensers to take advantage of refractive index variations. Fluorescence microscope. Considered one of the most versatile techniques of optical imaging, fluorescence microscopy uses a fluorescent substance e.

A fluorescent microscope uses a high-intensity illuminator which then excites the fluorophores in the sites of interest. As a result, the excited regions, in turn, emit light of a longer wavelength which makes it visible for observation.

Filters help produce the final image. Because it's more costly to conduct, fluorescence microscopy is usually reserved to important studies such as examining substances in low concentration. Practical applications of fluorescence microscopy include studies of porosity in ceramics, studies of semiconductors, and studies of nerve cells.

An ultraviolet microscope uses UV light to view specimens at a resolution that isn't possible with the common brightfield microscope. It utilizes UV optics, light sources, as well as cameras. Because of the shorter wavelengths of UV light nm , the image produced is clearer and more distinct at a magnification approximately double what is achieved by using only visible light nm. Confocal microscopy is regarded as a superior imaging technique that produces high-resolution, high-contrast images.

It uses fluorescence by focusing a laser over the specimen and collecting data from the emissions to reconstruct a final image. A common issue in viewing biological specimens via conventional light microscopy is glare captured from multiple focal planes producing light noise that can distort the image, especially if the specimen is thicker than the plane of focus. In confocal microscopy, spatial filtering is used to eliminate this glare by focusing light on a single point within a defined focal plane.

This produces impressively sharp images. Step 1: Connect the light microscope to a power source. If your microscope uses a mirror instead of an illuminator, you can skip this step.

Instead, find a place where natural light is easily accessible. Step 3: Mount your specimen onto the stage.