The two main optical components of the microscope which are adjusted and changed whilst in use are the objectives and the eyepieces. However, the other optical component which is sometimes overlooked, especially by beginners unaware of Koehler Illumination, is the microscope condenser.
To learn more about Koehler Illumination, please see this article from the Agar Scientific Blog;
On an upright microscope, the condenser is found beneath the stage (often referred to as a ‘sub-stage condenser’) and on an inverted configuration, it is found above the stage and the specimen. The condenser serves to focus the light waves from the light source to provide an even illumination and intensity across a specimen view. An integral part of the condenser is the aperture diaphragm and adjustments to the diaphragm, as well as varying the vertical position of the condenser, serve to change the contrast, depth-of-field and resolution of the image.
The history of the condenser
Robert Hooke (1635-1703) was an English scientist and natural philosopher. His interests and knowledge spanned many disciplines, from microscopy and palaeontology to the theory of gravity. In 1665, he published his book entitled ‘Micrographia’ which was to be a significant work in the field of microscopy and biology 1. Hooke was credited as being the first person to use the word ‘cell’ within the book. The microscope which Hooke used for his observations had a simple external condenser which consisted of a large globe filled with salt water and a plano-convex lens to focus the light on his specimens.
The modern achromatic objective lenses were developed in the 18th Century and such lenses corrected for chromatic aberration. When white light passes through a convex lens, it is split into its component wavelengths. The corrected (achromatic) lenses bring the red and blue wavelengths to approximately the same focal point as the green wavelength light. With such developments, it became apparent that there was a consequent need for an equal improvement in terms of condensers.
The first achromatic condenser was developed in 1837 by the French biologist Felix Dujardin (1801-1860) 2. However, improvements were subsequently made in the 1840’s by three prominent microscope makers; Andrew Ross, Hugh Powell and James Smith 2. In 1841, the Microscopical Society commissioned Hugh Powell to make a microscope for them and this particular one was fitted with an achromatic condenser which is thought to be the first ever to be produced in England 2. In contrast, the use of the condenser was not widespread in other European countries until 1870.
The Abbe condenser
Ernst Karl Abbe (1840-1905) was a German mathematician and physicist who not only co-founded Schott Glassworks in 1884, but also co-founded the Zeiss Optical Works with Carl Zeiss in 1866. Abbe was also the first person to define and describe numerical aperture (NA).
In 1870, Abbe invented the ‘Abbe Condenser’ which is still the most widely used design in light microscopy. This type of condenser consists of two or three lenses along with an associated diaphragm to control the amount of light with which the specimen is illuminated. The uppermost lens (on a three lens substage Abbe condenser) can be flipped into the light path which ensures that the light completely fills the field of view when using higher power objectives.
Condenser numerical aperture
Not only does the iris diaphragm control the light intensity, but adjusting this also determines the NA and size of the light cone from the condenser. Condensers have a mechanism similar to the focus wheels of a microscope which controls the vertical height of the condenser body. To realise the full optical potential and NA of the condenser and objectives, the condenser must be adjusted to the correct height for each objective used (see the above link to the article on Koehler Illumination). Whilst the Abbe condenser is adequate for most brightfield applications, its limitations come when using high magnification objectives as the maximum NA of a low-end Abbe condenser is around 0.6. It should be noted that to achieve an NA of greater than 0.95 requires the use of immersion oil between the top lens of the condenser and the underside of the specimen slide (conversely, ‘dry’ condensers have an NA of less than 0.95). Ideally, the condenser should have an NA which is equal to that of the highest power objective on the microscope.
As with the objectives, condensers are classified depending on their optical correction. Briefly, there are two types of aberrations for which condensers are corrected; spherical and chromatic (see above). Spherical aberrations occurs when light is focussed through a curved lens and the resulting light rays are spread along the optical axis instead of coming together at one focal point. For a more detailed description of aberrations, please see the article on objectives from the Agar Scientific Blog;
A simple two lens Abbe condenser has no optical corrections. An ‘aplanatic’ condenser is corrected for spherical aberration and may typically have a potential NA of up to 1.4. Achromatic condensers are corrected for chromatic aberration which brings the blue and red wavelength light to approximately the same focal point as the green wavelength. The highest level of correction is found in the aplanatic-achromatic condensers which are corrected for both spherical and chromatic aberrations.
In conclusion, the condenser plays a critical role in achieving the optimum resolution and highest potential NA from a microscope, therefore it is good practice to learn the correct set up for Koehler Illumination and not to overlook the importance of these optics!
- Bradbury, S 1967 The Evolution of the Microscope Pergamon Press, London.
AUTHOR: MARTIN WILSON