High-Resolution Video-Enhanced Differential Interference Contrast Light Microscopy

Abstract

Publisher Summary This chapter describes the basic concepts of differential interference contrast (DIC) image formation and analog and digital contrast enhancement, the practical aspects of component selection, alignment and specimen chambers for VE-DIC, and several test specimens for measuring microscope and video performance. A DIC microscope is a dual beam interferometer that uses a brightfield polarizing microscope containing polarizer, analyzer, and rotating stage. Contrast enhancement is produced by subtracting voltage from the video signal and amplifying the difference for each horizontal line scan of the video raster image. Increasing the analog contrast enhancement to the levels necessary to give distinct images of microtubules induces a new set of technical problems. Image quality becomes limited by noise from several sources, which is amplified as gain is increased. Fixed pattern noise could be eliminated from the video image by subtracting a stored background image, using a digital image processor operating at video rates. The background is obtained by defocusing the microscope until the microtubule image disappears. Achieving high-resolution, high-contrast VE-DIC images requires the highest performance of both the optical and video components of the video microscope system. Test specimens for microscope and video performance, such as diatom test slide, squamous cheek cell, and embedded skeletal muscle thin section, are also described in the chapter.