There are two sets of conjugate planes in Koehler Illumination. In the first set, the field diaphragm, objective front focal plane (specimen), the intermediate image plane and the retina are in conjugation with one another. In the second set, the filament, condenser focal plane, objective back focal plane and the iris of the eye are in conjugation. The latter is best viewed by removing the eyepice and inserting an eyepiece telescope or Bertrand lens.

The positions of conjugate planes in light microscopy. These planes are located where the light rays crossover. Note that there are two sets of conjugate focal planes in a light microscope adjusted for Koehler illumination.

The collector lens is located between the lamp and the field diaphragm. It gathers the light from the lamp, and magnifies and focuses an image of the filament at the front focal plane of the condenser (Set 2). This can be achieved by focusing the condenser using the condenser focus dial.

The field diaphragm is located in front of the condenser. It is used to adjust the illumination field reaching the specimen and should not exceed the capacity of the objective lens. Illuminating extraneous objects can cause light to scatter into the lens and cause glare. This will in turn, reduce contrast and resolution.

The condenser focuses light onto the specimen plane. This light then spreads from the specimen onto the objective lens (Set 1). The condenser also forms an image of the field diaphragm (Set 1).

The condenser diaphragm is adjusted so that the light achieves an angle that sufficiently fills the back focal plane of the objective lens (Set 2). This is important to achieve good resolution in the image. Removing the eyepiece allows you to view the back focal plane of the objective where an image of the condenser diaphragm appears. Adjusting the condenser levers focuses the image (Set 2).

The objective lens focuses the specimen image onto the intermediate focal plane (Set 1). This lens should also form an image of the filament at its back focal plane. (Set 2).

The ocular focuses the cone of light emerging from the image of the filament at the objective's back focal plane onto the eye's iris (Set 2).

The eye lens focuses the diverging rays of light from the ocular onto the retina where an image of the specimen is formed (Set 1). The filament, however, cannot be focused but is viewed as a field of light.