Darkfield microscopy is a technique to enhance contrast and make unstained samples visible under the microscope. The condenser for a darkfield microscope is designed to deliver a hollow intense cone of light with a greater N.A. than the objective N.A. Objectives with ah higher N.A. than 1.0 such as a 100x objective will require a funnel stop, or iris diaphragm. This technique will create a dark background around your sample making your unstained sample visible.
How To Achieve Darkfield
The simplest way to achieve darkfield is with a darkield stop. If you have a standard brightfield microscope and the condenser has a slot or a spot to accept a darkfield stop, you can convert it to darkfield. By simply inserting a darkfield stop into place you will achieve darkfield on all objectives but your 100x objective. Since a standard brighthfield 100x objective has a N.A. greater than 1.0 you will not achieve darkfield under this objective. You would need to purchase a darkfield funnel stop for this objective if available or purchase a replacement 100x objective with an iris diaphragm inside. By closing the iris diaphragm you can achieve a N.A. lower than 1.0.
The second way to achieve darkfield is with a dedicated darkfield oil condenser. This style condenser would only allow you to acheive darkfield. Using this condenser takes some time and patience to master as it involves applying a drop of oil to the top of the condenser lens. Before placing your slide on the stage of the microscope you apply a drop of oil to the top of the condenser lens. Once you have done so you then place your slide on the microscope and make sure that the oil on the top of the condenser is making contact with the underside of the slide. You will sill need to use oil on top of the cover slip while using the 100x oil objective.
Samples/applications that would require darkfield microscopy include : small aquatic organisms, blood cells, bacteria, alge, water samples, diamonds/precious stones, metal fractures.
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