Stereo microscopes are useful tools for examining specimens macroscopically and stereoscopically. These microscopes provide individual observation pathways for each eye, which gives the specimen some depth so that it appears as if you were examining it by eye. Stereo microscopes offer two main types of illumination: reflected illumination and transmitted illumination.
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The specimen type determines which illumination will work best. In many cases, you may need to use multiple methods. Each illumination type also has several observation methods, as shown in Table 1 below.
Table 1 – Commonly Used Observation Methods for Stereo Microscopes
| Observation Methods | |
|---|---|
| Reflected Illumination |
Oblique
Coaxial Polarized Fluorescence |
| Transmitted Illumination |
Brightfield
Darkfield Oblique Polarized |
Today, I’ll shine a light on transmitted illumination, so you can learn more about the observation methods within it.
4 Common Observation Methods for Transmitted Illumination
Now that we’ve covered the most commonly used observation methods for transmitted light, let’s explore their features and common applications.
Brightfield: Brightfield transmitted illumination is the most widely used method. Transmitted light is applied directly below the specimen. As light passes through the specimen, contrast is created by the attenuation of transmitted light through dense areas of the sample. If the sample isn’t stained and has no inherent color, then only small amounts of information can be obtained. In this case, it might be helpful to use other observation methods.
Darkfield: Darkfield transmitted illumination works well for samples that are unstained and transparent. The central light that typically passes through the specimen is blocked out, and only the oblique rays from every azimuth strike the specimen. These oblique rays are diffracted, reflected, and/or refracted by optical discontinuities. The result is a bright specimen on a black background.
Oblique Contrast: Oblique transmitted illumination also works well with unstained and transparent samples. This method enables you to adjust the angle of light to gain some contrast. It provides higher contrast compared to brightfield, but lower contrast compared to darkfield. With the oblique method, you can easily change the sample appearance to find the correct angle for illumination.
Polarized Illumination: Polarized transmitted illumination can be used on anisotropic specimens, which usually have multiple refractive indices (birefringent specimens). Polarization requires more equipment because the birefringent specimen must be placed between a polarizer and analyzer. The contrast comes from the interaction of plane-polarized light with the birefringent specimen, which will produce two separate wave components that are polarized in mutually perpendicular planes.
Zebrafish captured using brightfield observation
Medaka captured using oblique (left) and darkfield (right) observation methods
Expand the Scope of Observation
The ability to quickly and easily switch between observation methods can help you efficiently obtain information about your sample. Our LED transmitted-light stereo bases can help you switch between brightfield, oblique, darkfield, and polarized illumination—as well as different contrast methods—quickly and easily. Make sure to read our next blog post to see how!
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Brochure: Quad Position LED Transmitted Light Illumination Base
