Laboratory for Applied Microscopy Research-Light Microscopy, Marienburgstr. 23, Bullay, RLP-56859, Germany.
Microsc Microanal. 2012 Apr;18(2):343-52. doi: 10.1017/S1431927612000153. Epub 2012 Mar 2.
Variable phase dark-field contrast has been developed as an illumination technique in light microscopy, which promises significant improvements and a higher variability in imaging of several transparent specimens. In this method, a phase contrast image is optically superimposed on a dark-field image so that a partial image based on the principal zeroth-order maximum (phase contrast) interferes with an image that is based on the secondary maxima (dark field). The background brightness and character of the resulting image can be continuously modulated from a phase-contrast-dominated to a dark-field-dominated character. The condenser aperture diaphragm can be used for modulations of the image's appearance. Specimens can either be illuminated concentrically or obliquely (eccentrically) when parts of the illuminating light beams are covered and blocked. Moreover, a bright-field-like partial image can be added. In this way, the illumination can be optimally adjusted to the specific properties of the specimen. The techniques described can lead to improved visual information especially in biological specimens consisting of phase structures and additional light-absorbing or -reflecting components. Moreover, the specimen's three-dimensionality can be accentuated with improved clarity because the illuminating light beams associated with phase contrast and dark field run to the specimen at different angles of incidence.
可变相差暗场对比已被开发为一种在光学显微镜中的照明技术，它有望在对几个透明标本成像时带来显著的改进和更高的可变性。在这种方法中，一个相差图像被光学地叠加在暗场图像上，使得基于主零级最大值（相差）的部分图像与基于次级最大值（暗场）的图像相互干扰。所得图像的背景亮度和特征可以从相差主导连续调制为暗场主导。聚光器孔径光阑可用于调制图像的外观。当部分照明光束被覆盖和阻挡时，标本可以被同心或倾斜（偏心）照明。此外，可以添加一个类似于明场的部分图像。通过这种方式，可以根据标本的特定性质对照明进行最佳调整。所描述的技术可以提高视觉信息的质量，特别是在由相位结构和额外的吸光或反射成分组成的生物标本中。此外，由于与相差和暗场相关联的照明光束以不同的入射角照射到标本上，因此可以通过提高清晰度来突出标本的三维性。
