Conjugate Planes for Compound Microscopes
Click here to see why knowledge of conjugate planes is important.
Click here to learn what conjugate planes are.
Click here to learn about conjugate planes in Köhler Illumination.
Click here to learn about conjugate planes in Nelsonian Illumination.
Click here to learn how to view the conjugate planes.
This page describes the two sets of conjugate planes in the compound light microscope, for both Köhler Illumination and Nelsonian Illumination.
Why Do I Need to Know All of This?
On older microscopes equipped for Köhler illumination, an understanding of conjugate planes is absolutely essential for achieving proper Köhler setup, since this process involves complex positioning and centering of the lighting source, relative to the optics in the microscope. Köhler illumination focuses the lighting source at the conjugate illuminating (aperture) planes, and the field diaphragm at the conjugate image (field) planes, thereby illuminating the specimen under observation with perfectly defocused light which masks any non-uniformity present in the lamp. Because of this, Köhler Illumination provides consistent, high-contrast lighting for the specimen under observation, whereas Nelsonian Illumination is inherently plagued with field non-uniformity.
Later Köhler microscopes utilize an illumination source with the position and centering fixed in position, relative to the microscope optics, which greatly simplifies the setup of Köhler Illumination. This was in large part made possible by the availability of newer lamp technologies that provide filaments which are accurately positioned relative to the lamp base, allowing pre-centered Köhler designs to provide good results. A thorough understanding of conjugate planes is less necessary to set up these newer scopes, but all microscopists should nonetheless have a basic knowledge of conjugate planes.
What Are Conjugate Planes?
In microscopy, any planes in the optical system that are simultaneously in focus and appear to be superimposed are said to be “conjugate planes”. This means that an object on one plane will form a sharply focused image in all of its conjugate planes. In a properly aligned microscope, there are two distinct sets of alternating conjugate planes, which are known as the image (field) planes and the illumination (aperture) planes. The set of image planes are visible through the eyepieces, while the set of illumination planes can be viewed by pairing a Bertrand Lens with the eyepieces or by replacing one of the eyepieces with a phase telescope.
Any object placed into one of the conjugate image planes will appear to the observer to be sharply focused and superimposed with the specimen under observation. This is why a reticle positioned at the intermediate image plane within an eyepiece appears superimposed with the specimen, and also why the field diaphragm in a Köhler-equipped scope can be seen through the eyepieces during setup of Köhler illumination. In microscopes which use Nelsonian Illumination (rather than Köhler Illumination), this is also why any non-uniformity in the lamp filament can be seen in the specimen.
Description of the Conjugate Planes for Köhler-Equipped Microscopes
The two sets of conjugate planes are a bit different between a microscope equipped for Köhler Illumination, as compared to a microscope using Nelsonian Illumination. The conjugate planes for Köhler Illumination are described below.
Köhler Image Planes
The set of conjugate image (field) planes in a Köhler-equipped microscope consists of four planes where an object in the plane will appear in simultaneous focus with the specimen, as listed below. Any object placed into one of the conjugate image planes will appear sharply focused and overlaid with the specimen under observation. This is why a reticle positioned at the intermediate image plane within an eyepiece appears superimposed on the specimen, and why the field diaphragm in a Köhler-equipped scope can be seen during setup of Köhler illumination.
- Field diaphragm
- Specimen plane
- Intermediate image plane
- Observer’s retina / film or image sensor
In a microscope utilizing Köhler Illumination, the condenser focuses an image of the lamp filament onto the condenser aperture diaphragm. The set of conjugate illuminating (aperture) planes in a Köhler-equipped microscope consists of four planes which determine the angle, contrast, and intensity of the specimen illumination, as listed below. Because of the lighting uniformity inherent to Köhler Illumination, there is no necessity for a diffuser in the lighting path. In fact, many Köhler purists would argue that there should never be a diffuser in the lighting path. But Köhler zealots aside, there are in fact legitimate reasons why most modern Köhler-equipped scopes include a diffuser.
- Lamp filament or LED emitter
- Condenser aperture diaphragm
- Back focal plane of the objective
- Exit pupil of the eyepiece
Description of the Conjugate Planes for Microscopes Equipped for Nelsonian Illumination
The two sets of conjugate planes are a bit different between a microscope equipped for Köhler Illumination, as compared to a microscope using Nelsonian Illumination. The conjugate planes for Nelsonian Illumination are described below.
Nelsonian Image Planes
In a microscope utilizing Nelsonian Illumination, the condenser focuses an image of the lamp filament directly onto the specimen under observation. This of course adds the lamp filament to the set of conjugate image (field) planes in a Nelsonian-equipped microscope, which then consists of five planes where an object in the plane will appear in simultaneous focus with the specimen, as listed below. Any object placed into one of the conjugate image planes will appear sharply focused and overlaid with the specimen under observation. This is why a reticle positioned at the intermediate image plane within an eyepiece appears superimposed on the specimen, and why any non-uniformity present in the lamp filament will be visible in the resulting specimen image (even dust on the glass lamp bulb can be problematic). Because of the lighting non-uniformity inherent to Nelsonian Illumination, there should always be a diffuser in the lighting path to minimize this effect.
- Lamp filament of LED emitter
- Field diaphragm (if present)
- Specimen plane
- Intermediate image plane
- Observer’s retina / film or image sensor
Nelsonian Illuminating Planes
The set of conjugate illuminating (aperture) planes in a Nelsonian-equipped microscope consists of three planes which determine the angle, contrast, and intensity of the specimen illumination, as listed below:
- Condenser aperture diaphragm
- Back focal plane of the objective
- Exit pupil of the eyepiece
How To View the Conjugate Planes
The normal orthoscopic view provided by the eyepieces will directly show the conjugate image planes, allowing the operator to set up Köhler Illumination in modern, pre-centered scopes. Observation of the illuminating planes will allow the operator to set up old-school Köhler illumination and to align the phase annuli for phase-contrast microscopy (among other things). To view the conjugate illuminating planes in a microscope using either Köhler or Nelsonian Illumination, you must focus directly onto the light rays intersecting these conjugate planes. This can be done by using a Bertrand Lens in combination with the eyepieces (if your scope is so equipped), or by using a phase centering telescope in place of one of the eyepieces. The conoscopic image that this provides will allow you to directly view the rear focal plane of the objective.