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Dark Field Microscopy

11/7/16


Dark FieId Microscopy

In dark field microscopy the condenser is modified so that light hits the specimen from the side, and is reflected toward the objective when it strikes an object. The background is black and objects appear bright, often with false colors. Beautiful views of protozoa can be obtained in dark field mode. Details are well preserved at lower magnifications (e.g., 40x, 100x), however there is sufficient distortion at higher magnifications such that dark field is of limited use with the high dry lens (400x).

As with most specialized viewing modes, the effect is progressively degraded at higher magnifications as the thickness of the specimen increases. If the specimen is blurred in dark field even after cleaning the optics, it is probably too thick. Dark field mode is very useful in scanning a field for tiny unstained objects such as yeast cells or very small protozoa.

To use dark field illumination, adjust the condenser by inserting a dark field adapter or rotating the condenser turret to the dark field position. You may need to increase the illuminator intensity. You may also need to raise the condenser to just beneath the stage.

Adapting a bright field microscope

A dark field effect can be obtained on just about any microscope. The limiting factor is the intensity of light that can be directed to the specimen. At high magnifications, even a specially made condenser may be inadequate to provide reflected light of sufficient intensity. However with a compound microscope at low power and with any stereomicroscope (dissecting microscope) the effect can be obtained at very little expense.

Obtain an intense light source that produces a directed light, such as a fiber optic system, high intensity desk lamp with homemade reflector, or even a penlight (although a penlight will only work at the lowest magnifications). Simply block the light coming up through the specimen and shine the light source at a right angle to the light path coming into the objective. With some minor adjustments you will get a dark background with satisfactory illumination of all suspended particles, including bacteria, small eukaryotes,
dust, fibers, and just about everything.




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