Variable Phase Bright-Field Contrast—An Alternative Illumination Technique for Improved Imaging in Transparent Specimens

Abstract

In variable phase bright-field contrast, a bright-field image based on axial or concentric-peripheral light is optically superimposed with a phase-contrast image, so that typical details that are imminent in one or the other technique contribute to the resulting composite image. In particular, complex structured specimens consisting of high-density light absorbing details and additional low-density phase shifting components can be observed with improved clarity. As both partial images interfere with each other, fine details within thin specimens can be highlighted further by additional contrast effects based on interference. Haloing and shade-off are significantly reduced when compared with phase contrast carried out stand-alone. Our method is characterized by several technical means that are relevant for the high image quality that can be achieved: both illuminating light components associated with bright field and phase contrast are filtered at different colors and separated from each other so that they meet the specimen at different angles of incidence. The intensities of the phase-contrast- and bright-field-producing light can be selectively regulated so that the final image can be dominated by phase contrast or bright field, or be equalized. The condenser aperture diaphragm can be used for modulations of the image's appearance.