A method for preparing quick‐frozen, freeze‐substituted cells for transmission electron microscopy and immunocytochemistry
A method for preparing quick‐frozen, freeze‐substituted cells for transmission electron microscopy and immunocytochemistry
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一种用于透射电子显微镜和免疫细胞化学的速冻、冷冻替代细胞的制备方法
DOI：
10.1111/j.1365-2818.1985.tb02645.x
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发表时间：
1985
影响因子：
2
通讯作者：
H. Lee
中科院分区：
文献类型：
--
作者：
R. Nagele;M. Kosciuk;S. M. Wang;D. A. Spero;H. Lee
描述了一种快速冷冻、冷冻置换方法，该方法采用戊二醛和四氧化锇（OsO 4），采用与传统透射电子显微镜相当的“双重固定”方案。将培养的细胞在氟利昂22中快速冷冻，并在戊二醛的乙醇溶液中冷冻置换。将用于TEM的样本后固定在丙酮中的OsO 4中，包埋在Epon-Araldite中，并切片。该方法产生的超微结构保存与单独使用OsO 4作为冷冻置换固定剂的方法相当。但是，如果戊二醛单独用作冷冻置换固定剂，则可以对标本进行免疫细胞化学处理，而无需使用透化剂进行额外处理。
A quick‐freeze, freeze‐substitution method is described which employs glutaraldehyde as well as osmium tetroxide (OsO4) in a ‘double‐fixation’ protocol comparable to that used for conventional transmission electron microscopy. Cultured cells are quick‐frozen in Freon 22 and freeze‐substituted in an ethanolic solution of glutaraldehyde. Specimens destined for TEM are postfixed in OsO4 in acetone, embedded in Epon‐Araldite, and sectioned. This method yielded ultrastructural preservation which was comparable to that obtained from methods employing OsO4 alone as a freeze‐substitution fixative. However, if glutaraldehyde is used alone as a freeze‐substitution fixative, specimens can be processed for immunocytochemistry without additional treatment with permeabilizing agents.