Substage Condenser

The substage condenser contains a lens complex, an aperture diaphragm, and may also house prisms and filters used in contrast enhancement. The condenser can be focused to optimize the illumination of the specimen. The aperture diaphragm works independently of the field diaphragm to optimize contrast, depth of field, and resolution by altering the NA of the condenser lens (Fig. 2). The maximal NA of the condenser lens is usually engraved on its side. On some condensers, a swung-in lens can be used to alter the power of the condenser lens according to the choice of objective lens. Stopping down the aperture diaphragm will reduce the amount of light reaching the specimen, but unlike the field diaphragm, will not produce vignetting when the specimen is viewed through the microscope eyepieces.

Objective

Specimen

Condenser

Condenser front lens Aperture diapragm Filter wheel Focus

Field Diaphragm Light source

Fig. 2. The standard arrangement of light source, substage condenser, and specimen stage on a microscope designed for Köhler illumination. Two separate diaphragms, at the light source (field diaphragm) and on the condenser (aperture diaphragm), regulate the passage of light to the specimen.

As with objectives, the lenses in the condenser can be optically corrected in various ways, although this is not of primary importance for the resolution of the final image. Similarly, although it is possible to use immersion oil on virtually any high N.A. condenser lens, gains in image quality may be negligible.

3.4.1. Focusing the Condenser—"Köhler Illumination"

Bringing the condenser into focus is a simply a matter of bringing the edges of the field diaphragm into sharp focus (Fig. 3A).

1. Focus the objective lens so that the image of the specimen is sharp through the eyepiece.

2. Where the condenser has a range of front lenses and, particularly, if it has a range differential interference contrast (DIC) prisms, check that the NA marked on the condenser lens and on the DIC adjustment collar matches that of the objective.

3. Close the field diaphragm so that its edges are in view, and center the circle of illumination with adjuster screws on the substage condenser.

4. Adjust the focus of the condenser such that the edges of the field diaphragm are sharp.

5. Open the field diaphragm to illuminate the field of interest, but to minimize the amount of stray light. The edges of the field diaphragm should remain just inside the field of view.

6. Repeat this procedure every time an objective is changed.

3.4.2. Aperture Diaphragm Adjustment

This adjustment matches the NA of the condenser to that of the objective, optimizing the cone of light that passes through the specimen (Fig. 3B).

Objective

Specimen

Condenser

Condenser front lens Aperture diapragm Filter wheel Focus

Fig. 2. The standard arrangement of light source, substage condenser, and specimen stage on a microscope designed for Köhler illumination. Two separate diaphragms, at the light source (field diaphragm) and on the condenser (aperture diaphragm), regulate the passage of light to the specimen.

A Field diaphragm

A Field diaphragm

Fig. 3. The steps involved in focusing the condenser, optimizing the field diaphragm (A) and the condenser aperture diaphragm (B) (see Subheadings 3.4.1. and 3.4.2.). Schematic views through the eyepiece for each step are shown below.

1. Focus the condenser as above (see Subheading 3.4.1.).

2. Remove the eyepiece, and ensure that the field diaphragm is sufficiently open to fully illuminate the field of view.

3. Close the aperture diaphragm on the substage condenser so that its radius is 70-80% of the illuminated field (the "exit pupil" of the objective). Screening off the outer 20-30% of the exit pupil gives an approximate match between the NAs of condenser lens and objective lens.

4. Replace the eyepiece.

5. Repeat this procedure every time an objective is changed.

The 20-30% screening of the exit pupil is a "rule of thumb," and the final adjustment of the aperture diaphragm alters image quality in various ways. As the aperture diaphragm is closed, resolution decreases, but both contrast and the depth of field increase (see Note 7). As stated above, there should be no vignetting when the aperture diaphragm is adjusted, only a uniform variation in lighting intensity.

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Responses

  • arnaldo
    How to regulate the substage condenser of a microscope?
    1 year ago
  • louis
    How can a substage condenser be regulated?
    1 year ago
  • mareta
    What is the use of a sub stage condenser?
    10 months ago
  • Alistair
    How can a substage condenser be regulated on a microscope?
    7 months ago
  • roisin
    What is the use of sub stage condenser on a light microscope?
    2 months ago
  • lukasz
    How do i regulate substage condenser and its function?
    23 days ago
  • Orazio
    Where is substage condenser located, how can it be regulated and its function?
    20 days ago

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