免疫电镜下用柠檬酸酐提取抗原的新方法:在II型肺泡上皮细胞中定位表面活性剂前蛋白C (proc -C)。

W Dai, S Sato, M Ishizaki, K Wakamatsu, S Namimatsu, Y Sugisaki, M Ghazizadeh
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引用次数: 0

摘要

为了建立一种改进超微结构形态的预包埋免疫电镜方法,我们研究了一种使用柠檬酸酐的抗原回收(AR)方法,并比较了戊二醛固定与常规多聚甲醛固定在预包埋免疫电镜方法中的效果,并参考了表面活性剂相关的前蛋白C (pro -C)在肺中的定位。经戊二醛固定的组织在0.05%柠檬酸酸酐溶液中(pH 7.4), 98℃下浸泡60 min后进行免疫染色。常规多聚甲醛固定的预包埋免疫电镜法发现,II型肺泡上皮细胞中散在分布着prospc阳性产物。在未采用AR法的戊二醛固定组织中,未检测到普洛斯- c产物,而在柠檬酸酐中进行AR后,普洛斯- c阳性产物特异性分布于粗内质网膜、高尔基复合体膜、多泡体和亲锇层状体中。prob - c阳性产物在肺泡内也呈格状结构。因此,本AR方法提供了戊二醛固定组织的成功预包埋免疫电镜。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
A new antigen retrieval method using citraconic anhydride for immunoelectron microscopy: localization of surfactant pro-protein C (proSP-C) in the type II alveolar epithelial cells.

In the attempt to develop a pre-embedding immunoelectron microscopic method with improved ultrastructral morphology, we examined an antigen retrieval (AR) method using citraconic anhydride, and compared the effects of glutaraldehyde fixation with routine paraformaldehyde fixation in the pre-embedding immunoelectron microscopic method with reference to the localization of surfactant-associated pro-protein C (proSP-C) in the lung. The glutaraldehyde-fixed tissues were immunostained after AR in 0.05% citraconic anhydride solution, pH 7.4, at 98 degrees C for 60 min. In routine pre-embedding immunoelectron microscopic method using paraformaldehyde fixation, proSP-C positive products were distributed sporadically in the type II alveolar epithelial cells. In glutaraldehyde-fixed tissues without the AR method, proSP-C products were not detected, however after AR in citraconic anhydride proSP-C positive products were distributed specifically, in rough endoplasmic reticulum membranes, Golgi complex membranes, multivesicular bodies and osmiophilic lamellar bodies. The positive proSP-C products also showed lattice-like structures in the alveoli. Thus, the present AR method provides successful pre-embedding immunoelectron microscopy of glutaraldehyde-fixed tissues.

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