THE STRUCTURE OF CYTOPLASM IN DIRECTLY FROZEN CULTURED-CELLS .1. FILAMENTOUS MESHWORKS AND THE CYTOPLASMIC GROUND SUBSTANCE
THE STRUCTURE OF CYTOPLASM IN DIRECTLY FROZEN CULTURED-CELLS .1. FILAMENTOUS MESHWORKS AND THE CYTOPLASMIC GROUND SUBSTANCE
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DOI：
10.1083/jcb.99.5.1655
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发表时间：
1984-01-01
影响因子：
7.8
通讯作者：
REESE, TS
中科院分区：
文献类型：
--
作者：
BRIDGMAN, PC;REESE, TS
将培养的非洲爪哇胚胎成纤维细胞或上皮细胞直接冷冻，用改进的方法冷冻置换，然后进行临界点干燥，作为整体，或包埋，薄片。在这些细胞的这个区域，没有冰晶伪影，细胞质由密集的、高度相互连接的丝网组成，嵌入细小的颗粒基质中。将直接冷冻的完整细胞中的网状结构与冷冻前裂解(物理上、用洗涤剂或非磷脂)或用戊二醛固定的细胞中的网状结构进行比较。尽管裂解细胞中的细丝数量较少，但它们的整体组织结构与完整细胞中的组织结构相同。冷冻前用戊二醛固定，会根据固定缓冲液的渗透压不同程度地扭曲网状结构，也会遮盖直接冷冻细胞中明显的颗粒状基质。在最佳的制备方法下，这些直接冷冻的细胞的细胞质由离散的相互交织的细丝和容易提取的颗粒基质组成，这些细丝由许多细丝相连，颗粒基质可能代表细胞质蛋白质的聚集。
Cultured fibroblasts or epithelial cells derived from Xenopus laevis embryos were directly frozen, freeze-substituted by an improved method, and then either critical-point-dried and viewed as whole mounts, or embedded and thin sectioned. In this regions of these cells, where ice crystal artifacts are absent, the cytoplasm consisted of a dense, highly interconnected meshwork of filaments, embedded in a finely granular ground substance. The meshwork in directly frozen, intact cells was compared with that in cells that were lysed (physically, with detergents, or with filipin), or fixed with glutaraldehyde before freezing. Although filaments tended to be less numerous in lysed cells, their overall organizations was the same as that in intact cells. Fixation with glutaraldehyde before freezing distorted the meshwork to variable degrees depending on the osmolarity of the fixation buffer, and also obscured the granular ground substance which is obvious in directly frozen cells. With optimal preparative methods, the cytoplasm of these directly frozen cells is shown to consist of a cytoskeleton composed of discrete interwoven filaments interconnected by numerous finer filaments and a readily extractable granular matrix which presumably represents aggregations of cytoplasmic proteins.