Ling Erjun, Shirai Koji, Kanekatsu Rensuke, Kiguchi Kenji
Laboratory of Silkworm Physiology, Department of Applied Biology, Faculty of Textile Science and Technology, Shinshu University, Ueda, 386-8567 Nagano, Japan.
Histochem Cell Biol. 2003 Dec;120(6):505-11. doi: 10.1007/s00418-003-0592-6. Epub 2003 Nov 11.
Circulating hemocytes of the silkworm can be classified by fluorescence microscopy following staining with acridine orange and propidium iodide. Based on their fluorescence characteristics, three groups of circulating hemocytes can be distinguished. The first group, granulocytes and spherulocytes, is positive for acridine orange and contain bright green fluorescent granules when observed by fluorescence microscopy. In granulocytes, these green granules are heterogeneous and relatively small. In contrast, in spherulocytes, the green granules appear more homogenous and larger. The second group of hemocytes consists of prohemocytes and plasmatocytes. These cells appear faint green following staining with acridine orange and do not contain any green fluorescent granules in the cytoplasm. Prohemocytes are round, and their nuclei are dark and clear within a background of faint green fluorescence. Inside the nucleus there are one or two small bright green fluorescent bodies. Plasmatocytes are irregularly shaped and their nuclei are invisible. Oenocytoids belong to the third group, and their nuclei are positive for propidium iodide. Therefore, all five types of circulating hemocytes of the silkworm, including many peculiar ones that are difficult to identify by light microscopy, can now be easily classified by fluorescence microscopy following staining with acridine orange and propidium iodide. In addition, we show that hemocytes positive for acridine orange and propidium iodide are in fact living cells based on assays for hemocyte composition, phagocytosis, and mitochondrial enzyme activity.
家蚕的循环血细胞在用吖啶橙和碘化丙啶染色后，可通过荧光显微镜进行分类。根据它们的荧光特性，可以区分出三组循环血细胞。第一组，粒细胞和球血细胞，对吖啶橙呈阳性，在荧光显微镜下观察时含有亮绿色荧光颗粒。在粒细胞中，这些绿色颗粒是异质的且相对较小。相比之下，在球血细胞中，绿色颗粒显得更均匀且更大。第二组血细胞由原血细胞和浆血细胞组成。这些细胞在用吖啶橙染色后呈淡绿色，细胞质中不含任何绿色荧光颗粒。原血细胞呈圆形，在淡绿色荧光背景下，其细胞核黑暗且清晰。细胞核内有一两个小的亮绿色荧光体。浆血细胞形状不规则，其细胞核不可见。类绛色细胞属于第三组，其细胞核对碘化丙啶呈阳性。因此，家蚕所有五种类型的循环血细胞，包括许多通过光学显微镜难以鉴定的特殊类型，现在在用吖啶橙和碘化丙啶染色后，通过荧光显微镜可以很容易地进行分类。此外，基于血细胞组成、吞噬作用和线粒体酶活性的测定，我们表明对吖啶橙和碘化丙啶呈阳性的血细胞实际上是活细胞。
