Contact & Support
Header Image

Confocal Microscopy Basics

Basic Principle of Confocal Microscopes

In the confocal microscope all structures out of focus are suppressed at image formation.

This is obtained by an arrangement of diaphragms which, at optically conjugated points of the path of rays, act as a point light source and as a point detector respectively.

Rays from out-of-focus are suppressed by the detection pinhole.

The depth of the focal plane is, besides the wavelength of light, determined in particular by the numerical aperture of the objective used and the diameter of the diaphragm.

At a wider detection pinhole the confocal effect can be reduced.

To obtain a full image, the image point is moved across the specimen by mirror scanners.

The emitted/reflected light passing through the detector pinhole is transformed into electrical signals by a photomultiplier and displayed on a computer monitor screen.

Advantages of Confocal Microscopes

Major improvements offered by a confocal microscope over the performance of a conventional microscope may be summarized as follows:

  • Light rays from outside the focal plane will not be recorded.
  • Defocussing does not create blurring, but gradually cuts out parts of the object as they move away from the focal plane. Thus, these parts become darker and eventually disappear. This feature is called optical sectioning.
  • True, three-dimensional data sets can be recorded.
  • Scanning the object in x/y-direction as well as in z-direction (along the optical axis) allows viewing the objects from all sides.
  • Due to the small dimension of the illuminating light spot in the focal plane, stray light is minimized.
  • By image processing, many slices can be superimposed, giving an extended focus image which can only be achived in conventional microscopy by reduction of the aperture and thus sacrificing resolution