Off-Axis Illumination Transitions
The true versatility of the unique MIC-D digital microscope design becomes apparent with oblique and darkfield illumination techniques made possible by off-axis translation of the illuminator head and condenser assembly. This feature enables the microscopist to enhance contrast in specimens that would otherwise remain invisible (or nearly so) in brightfield illumination. The interactive tutorial explores illumination transitions, starting at brightfield, moving through the varying degrees of oblique, and finally reaching darkfield.
The tutorial initializes with a randomly selected specimen appearing in a miniature version of the MIC-D interface software Live Image screen. A cutaway diagram of the MIC-D microscope illuminator, stage, and objective appear in the right-hand side of the tutorial window. Initially, the specimen presented in the Live Image screen is illuminated with on-axis brightfield lighting, which is indicated above the Illumination Mode slider. To vary specimen illumination by tilting the illumination head off-axis, translate the Illumination Mode slider to the right. As the slider is moved, the illumination mode indicator above the slider will change from Brightfield to Oblique and then to Darkfield. Simultaneously, the contrast of the specimen appearing in the Live Image will change to reflect how the specimen appears when the illuminator is tilted. In order to select another specimen, use the Choose A Specimen pull-down menu.
In oblique illumination, direct light from the condenser light cone is restricted to a single azimuth, striking the specimen from only one direction rather than bathing it with an even distribution of light through a well-defined numerical aperture. The net effect is to reveal details in pseudo-relief to produce a shadowed effect on the side opposite the light source and bright specimen highlights on the side nearest the illuminator. In order to achieve oblique or off-axis illumination with the MIC-D digital microscope, the illuminator head is shifted by moving the rotation arm a few degrees away from the upright (brightfield) position.
The useful range of motion for producing oblique illumination with the rotation arm is between 2-3 and 15 degrees from the vertical (optical) axis of the MIC-D digital microscope. In addition, the off-axis angle of specimen illumination can also be modulated or fine-tuned by rotating the illumination head itself (while the arm is held stationary) on a pivot that attaches the head to the rotation arm. The illumination head can be rotated on the pivot to a maximum angle of 10 degrees. Because there are two variables that can be introduced when producing off-axis illumination with the MIC-D, rotation of the main illumination arm and independent rotation of the attached illumination head, the number of possibilities for varying the illumination angle is almost limitless.
In traditional optical microscopy, darkfield illumination requires blocking the central light that ordinarily passes through and around (surrounding) the specimen, allowing only oblique rays from every azimuth to "strike" the specimen mounted on a microscope slide. When the rotation arm of the MIC-D digital microscope is positioned at a distance greater than 15 degrees from the central (optical) axis, this instrument is capable of imaging specimens by a mechanism that produces results similar to those observed in true darkfield illumination.
Matthew J. Parry-Hill and Michael W. Davidson - National High Magnetic Field Laboratory, 1800 East Paul Dirac Dr., The Florida State University, Tallahassee, Florida, 32310.
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