Laboratory Instrument: Microscope

Microorganisms are too small to see unaided. In order to "see" them we need to magnify an image or collect a large number of the microbes in one place. We can observe a large number of organisms when we look at colonies on a plate. We look at individual organisms using a microscope. Standard microscopes amplify the image through two lenses. One lens is in the eyepiece and the other is in the objective. To calculate the overall magnification one multiplies the magnifying power of each lens together. Eyepieces usually contain lenses that amplify ten times. Objectives vary in a magnification and most microscopes contain several objectives from which the microscopist may choose.

(10X eyepiece lens)(40X objective lens) = 400X total amplification

  • Question
  • Answer

The top magnification for a light microscope is 1000X. What objective is used for a 1000X magnification?

100X
(1000X = 10X eyepiece lens * 100X objective lens)

 

Working distance is the space between the objective and the specimen or slide. The focal distance increases with the working distance. The magnification is inversely related to the working distance. Therefore the 100X objective would have the smallest working distance.

The top magnification for a light microscope is about 1000X. Above this, the limits of the optical system produce a fuzzy image. This is related to the resolving power, or resolution, of the microscope. The resolving power or resolution of the microscope is the ability of the microscope or optical instrument to distinguish between two very close objects.

Resolving power = wavelength of light used/(Numerical aperture of the objective aperture + Numerical aperture of the sub-stage condenser)

The numerical aperture is a measure of the size of the cone of light that may be gathered by the objective. The largest cone of light is needed for the highest magnification. With low magnification the working distance is increased, less definition is required, and the numerical aperture setting is lower. The microscopist can control the light as it passes through the sub-stage condenser and may use this to change the numerical aperture.

For more information about microscopes and how they work see the [Microscope Operating Guide].

Below are several links to the Olympus Microscope Resource Center, a wonderful microscope website, for you to explore. If you open these links you will open a new window that is not connected to the Microbiology eGuide site.

This site reviews the numerical aperture and the light cone entering the objective.
http://www.olympusmicro.com/primer/java/microscopy/immersion/index.html

This site reviews the substage condenser function.
http://www.olympusmicro.com/primer/java/kohler/contrast/index.html

This site reviews how the substage condenser controls the light cone that illuminates the specimen.
http://www.olympusmicro.com/primer/java/kohler/condenseraperture/index.html

On to Operating Guide

On to Instrument Diagram

On to Microscope Parts Self Quiz

Take the Microscope Quiz

Take the Light Path Quiz