All of us are quite familiar with the appearance and visibility of stars on a dark night, this despite their vast distances from the Earth. Stars can be readily observed at night primarily because of the stark contrast between their faint light and the black sky. Stars are shining both night and day, but they are invisible during the day because the overwhelming brightness of the sun blots out the faint light from the stars, rendering them invisible. During a total solar eclipse, the moon moves between the Earth and the sun blocking out the light of the sun and the stars can now be seen even though it is daytime. In short, the visibility of the faint star light is enormously enhanced against a dark background. This principle is applied in darkfield, also called dark ground, microscopy, and a simple and popular method for making unstained transparent specimens clearly visible. Such objects often have refractive indices very close in value to that of their surroundings and are difficult to image in conventional brightfield microscopy. For instance, many small aquatic organisms have a refractive index ranging from 1.2 to 1.4, resulting in a negligible optical difference from the surrounding aqueous medium. These are ideal candidates for darkfield illumination.

Darkfield illumination requires blocking out of the central light which ordinarily passes through and around, surrounding the specimen, allowing only oblique rays from every azimuth to strike the specimen mounted on the microscope slide. The top lens of a simple darkfield condenser is spherically concave, allowing light rays emerging from the surface in all azimuths to form an inverted hollow cone of light with an apex centered in the specimen plane. If no specimen is present and the numerical aperture of the condenser is greater than that of the objective, the oblique rays cross and all such rays will miss entering the objective because of their obliquity. The field of view will appear dark. The darkfield condenser or objective pair is a high-numerical aperture arrangement that represents darkfield microscopy in its most sophisticated configuration, which will be discussed in detail below. The objective contains an internal iris diaphragm that serves to reduce the numerical aperture of the objective to a value below that of the inverted hollow light cone emitted by the condenser. The cardioid condenser is a reflecting darkfield design that relies on internal mirrors to project an aberration-free cone of light onto the specimen plane. When a specimen is positioned on the slide, especially an unstained, non-light absorbing specimen, the oblique rays cross the specimen and are diffracted, reflected, and/or refracted by optical discontinuities such as the cell membrane, nucleus, and internal organelles allowing these faint rays to enter the objective. The specimen can then be seen bright on an otherwise black background. In terms of Fourier optics, darkfield illumination removes the zeroth order, unscattered light, from the diffraction pattern formed at the rear focal plane of the objective. This result in an image formed exclusively from higher order diffraction intensities scattered by the specimen. The advantage of darkfield illumination is that you can notice details that are normally not resolved by the microscopes objective. You can not see the actual detail but because it reflects light you can see it. A good analogy here is that of dust in a room. In a well lit room you do not see the very small dust particles. Nevertheless, if the lights go out, a beam of light from an acute angle makes these same particles visible. Besides the optical advantages darkfield illumination is very beautiful and gives an almost science fiction like image.