ABSTRACT
The goal of the illumination portion of the microscope is to provide bright and even illumination across the field of view while not illuminating any areas outside of the field of view. This reduces scattered light and photodamage. To accomplish this, the microscope is aligned such that the illumination set of planes are separate from the set of planes that carry the specimen information. The real advantage of fluorescence microscopy is its ability to detect fluorescent objects because a fluorescent object is very bright relative to the background. As we know, fluorescence emitted from the specimen is radiated in all directions. A good rule of thumb is that if the objective aperture is doubled in size, approximately four times more fluorescence light can be gathered. Oil immersion also eliminates the loss of light caused by light reflection on the surfaces. Lastly, none of the optical components of the microscope should be autofluorescent.
