Žižka Zdeněk, Gabriel Jiří
Institute of Microbiology, Academy of Sciences CR, Vídeňská 1083, 142 20, Prague 4-Krč, Czech Republic.
Folia Microbiol (Praha). 2015 Nov;60(6):545-50. doi: 10.1007/s12223-015-0397-8. Epub 2015 May 21.
Polarization and positive phase contrast microscope were concomitantly used in the study of the internal structure of microbial cells. Positive phase contrast allowed us to view even the fine cell structure with a refractive index approaching that of the surrounding environment, e.g., the cytoplasm, and transferred the invisible phase image to a visible amplitude image. With polarization microscopy, crossed polarizing filters together with compensators and a rotary stage showed the birefringence of different cell structures. Material containing algae was collected in ponds in Sýkořice and Zbečno villages (Křivoklát region). The objects were studied in laboratory microscopes LOMO MIN-8 Sankt Petersburg and Polmi A Carl Zeiss Jena fitted with special optics for positive phase contrast, polarizers, analyzers, compensators, rotary stages, and digital SLR camera Nikon D 70 for image capture. Anisotropic granules were found in the cells of flagellates of the order Euglenales, in green algae of the orders Chlorococcales and Chlorellales, and in desmid algae of the order Desmidiales. The cell walls of filamentous algae of the orders Zygnematales and Ulotrichales were found to exhibit significant birefringence; in addition, relatively small amounts of small granules were found in the cytoplasm. A typical shape-related birefringence of the cylindrical walls and the septa between the cells differed in intensity, which was especially apparent when using a Zeiss compensator RI-c during its successive double setting. In conclusion, the anisotropic granules found in the investigated algae mostly showed strong birefringence and varied in number, size, and location of the cells. Representatives of the order Chlorococcales contained the highest number of granules per cell, and the size of these granules was almost double than that of the other monitored microorganisms. Very strong birefringence was exhibited by cell walls of filamentous algae; it differed in the intensity between the cylindrical peripheral wall and the partitions between the cells. Positive phase contrast enabled us to study the morphological relationship of various fine structures in the cell (poorly visible in conventional microscope) to anisotropic structures that have been well defined by polarization microscopy.
在微生物细胞内部结构的研究中，同时使用了偏光显微镜和正相衬显微镜。正相衬显微镜使我们能够观察到甚至是那些折射率与周围环境相近的精细细胞结构，比如细胞质，并将不可见的相位图像转化为可见的振幅图像。利用偏光显微镜，交叉偏振滤光片与补偿器以及旋转载物台显示出不同细胞结构的双折射现象。含有藻类的材料采集自锡科里采村和兹贝乔村（克里沃克拉特地区）的池塘。这些样本在装有用于正相衬的特殊光学器件、偏振器、检偏器、补偿器、旋转载物台的圣彼得堡LOMO MIN - 8显微镜和耶拿卡尔·蔡司Polmi A显微镜以及用于图像采集的尼康D 70数码单反相机上进行研究。在裸藻目鞭毛虫的细胞、绿球藻目和小球藻目的绿藻以及鼓藻目的鼓藻中发现了各向异性颗粒。双星藻目和丝藻目的丝状藻细胞壁表现出显著的双折射现象；此外，在细胞质中发现了相对少量的小颗粒。圆柱状细胞壁和细胞间隔膜典型的形状相关双折射在强度上有所不同，在连续两次设置过程中使用蔡司补偿器RI - c时尤其明显。总之，在所研究的藻类中发现的各向异性颗粒大多表现出强烈的双折射现象，并且在细胞中的数量、大小和位置各不相同。绿球藻目的代表细胞中每个细胞含有的颗粒数量最多，这些颗粒的大小几乎是其他被监测微生物颗粒大小的两倍。丝状藻的细胞壁表现出非常强的双折射现象；圆柱状外周壁和细胞间隔膜之间的双折射强度不同。正相衬使我们能够研究细胞中各种精细结构（在传统显微镜下难以看清）与通过偏光显微镜已明确界定的各向异性结构之间的形态关系。
