It enables the increasing of micrograph contrast and the reconstruction of 3D images.

Confocal microscopy offers many advantages in optical imaging. These include greater control of the depth of field, the removal of degradation due to out-of-focus information, and the capability to gather sectional information from specimens that are quite thick. Today, confocal microscopy is being used widely in various fields of science and industrial applications as well.

Basic concept behind confocal microscopy

How does a confocal microscope work?

In contrast to conventional microscopy which makes use of a flood of light to illuminate the specimen, confocal microscopy makes use of point illumination.

This is done by using a pinhole whereby the light is made to pass through. As a result, only the light that is covered by the illumination plane is detected, getting rid of the out-of-focus information.

The concept of confocal microscopy is actually attributed to Marvin Minsky who patented it in 1957.

They are confocal laser scanning microscopes, spinning disk confocal microscopes, and Programmable Array Microscopes.

Confocal laser scanning microscopes are used to obtain in-focus images of thick specimens. This is done by virtually cutting the specimens into sections. These sections are then reconstructed point by point using a computer. The end result is a very accurate image of the whole specimen, despite its thick nature.

Spinning disk confocal microscopes are also called Nipkow disk microscopes. Developed by Paul Nipkow in 1884, these disks allow for greater transmission of light. However, they less focus than the laser scanning microscopes discussed above. There are also other types of spinning disk confocal microscopes but the Nipkow disks are the most popular and most widely used type. The sector that uses spinning disk confocal microscopes over laser scanning microscopes are usually scientists who work with 4D imaging of living cells. This is because spinning disk confocal microscopes yield video rate imaging.

The third type, Programmable Array Microscopes, or PAMs, result in much better image quality as compared to spinning disk confocal microscopes. On the downside, they result in low frame imaging rate. This means that they can only capture 3 frames per second, which is not good enough for certain applications.

Individuals who desire to learn more about confocal microscopy can find a host of tutorials on the Internet. These cover the various techniques and tools in great detail.

Microscopy is a technique that produces magnified images of small details.

It is widely used in biotechnology, material sciences, nanotechnology and precision measurements.

See also

• Optical and Electron Microscopy
• Bright Field Optical Microscopy
• Dark Field Microscopy

This can be carried out by standard light microscopy, transmission electron microscopy or by scanning a fine beam over the sample as in confocal microscopy and scanning electron microscopy.

Scanning probe microscopy involves the interaction of the scanning probe with the surface or object of interest.

Optical or light microscopy involves the passing of visible light transmitted through or reflected from the sample through a single or multiple lenses to allow a magnified view of the sample. The resultant image can be detected visually or imaged digitally or on a photographic plate. The basic microscope is a simple staging area and support along with a single lens or a system of lenses, including attachments.

The technique of optical microscopy can only image dark or strongly refracting objects effectively. Diffraction will limit the resolution to 0.2 micrometers. Out of focus light from places outside the focal plane reduces the image clarity.

Live cells, for example, lack sufficient contrast to be studied as internal structures are often colorless and transparent. The best way to see these structures is to stain the different structures with selective dyes. This involves the killing of the cells and fixing the sample. Staining is not perfect and can produce artifacts, structural details that are caused by the processing of the specimen.

Oblique illumination has the illumination coming in from the side and it gives a three dimensional appearance, highlighting otherwise invisible features. A more recent technique is based on a method called Hoffmann’s modulation contrast, a system found on inverted microscopes for use in cell cultures. It suffers from some of the same limitations as bright field microscopy but may highlight invisible structures.

Dark Field Microscopy can dramatically improve the visibility of transparent objects.