Tamaki H, Yamashina S
Department of Anatomy, Kitasato University School of Medicine, Japan.
J Histochem Cytochem. 1994 Sep;42(9):1285-93. doi: 10.1177/42.9.8064136.
We established an improved method for post-embedding cytochemistry by which highly specific cytochemical reactions on excellent cellular ultrastructures are possible. The method is a combination of post-fixation in potassium ferrocyanide-reduced OsO4 and embedment in acrylic-based LR White resin. It permits both immuno- and lectin-gold cytochemistry with fine ultrastructures comparable to those obtained by conventionally osmicated and epoxy-embedded tissues. Fixation with reduced osmium appeared to contribute to the preservation of immunoreactivity and membranous structures. By this method, the immunocytochemical localization of secretory proteins (amylase and chymotrypsinogen), actin filaments by polyclonal antibodies, 105 KD Golgi-associated protein by a monoclonal antibody (GF-1), and binding sites for gold-labeled lectin could be demonstrated. Also possible was multiple staining with enzyme cytochemistry of thiamine pyrophosphatase and immunocytochemistry of GF-1 and anti-amylase antibodies. This multiple staining made possible partial characterization of the trans-Golgi network in parotid acinar cells.
我们建立了一种改进的包埋后细胞化学方法，通过该方法可以在优异的细胞超微结构上进行高度特异性的细胞化学反应。该方法是在亚铁氰化钾还原的OsO4中进行后固定，并嵌入丙烯酸基LR White树脂。它允许进行免疫和凝集素金细胞化学，其精细的超微结构与通过传统锇固定和环氧树脂包埋的组织所获得的超微结构相当。用还原锇固定似乎有助于免疫反应性和膜结构的保存。通过这种方法，可以证明分泌蛋白（淀粉酶和胰凝乳蛋白酶原）的免疫细胞化学定位、用多克隆抗体对肌动蛋白丝的定位、用单克隆抗体（GF-1）对105 KD高尔基体相关蛋白的定位以及金标记凝集素的结合位点。用硫胺焦磷酸酶的酶细胞化学和GF-1及抗淀粉酶抗体的免疫细胞化学进行多重染色也是可能的。这种多重染色使得对腮腺腺泡细胞中反式高尔基体网络进行部分表征成为可能。
