What is darkfield illumination?

Darkfield Illumination

Darkfield illumination is a microscopy technique used to enhance the contrast in unstained samples. Unlike traditional brightfield illumination, which illuminates the sample directly, darkfield illumination directs light toward the specimen at oblique angles. This method ensures that only the light scattered by the specimen enters the microscope’s objective, making the specimen appear bright against a dark background, thus the name 'darkfield'.

How It Works

In darkfield microscopy, a special condenser is used that blocks direct light from entering the objective lens. This condenser has an opaque disc in its center, which prevents unperturbed light from illuminating the sample directly. Instead, light is directed to the sides of this disc, and then it is refracted or diffracted into the specimen. The only light that enters the objective and forms the image is the light scattered by the sample, making it appear brightly lit against a dark, almost completely black, background.

Applications

• Observing living cells and organisms that are invisible in other illumination modes.
• Studying the motility of bacteria and other microorganisms.
• Viewing the details in transparent specimens such as pond water organisms.
• Examining the surface features of materials or the structural details in minerals.

Advantages

• Enhances the visibility of transparent and unstained samples.
• Allows for the observation of living specimens without the need for staining.
• Provides excellent contrast, highlighting details that are not visible with brightfield illumination.

Disadvantages

• Lower light levels mean that darkfield images can be grainier than those from brightfield illumination.
• Not suitable for observing very thick specimens as the technique maximizes detail in thin samples.
• Requires a specialized condenser, which can be an additional expense.