Hearn S A, Silver M M, Sholdice J A
J Histochem Cytochem. 1985 Dec;33(12):1212-8. doi: 10.1177/33.12.3934258.
Previous studies have found that immunoglobulin cannot be immunolabeled in tissues prepared for electron microscopy by usual methods. To test this conclusion, we used a protein A-gold postembedding immunolabeling method on tissues that were fixed in glutaraldehyde, post-fixed in osmium tetroxide, and embedded in epoxy resin; sections were pretreated with sodium metaperiodate. A variety of common fixation protocols were also used and the most suitable conditions for immunolabeling were determined. This technique permitted the ultrastructural localization of immunoglobulin light chains in optimally preserved and contrasted plasma cells from human tonsil, lymph nodes, plasmacytomas, and a renal biopsy. We were able to demonstrate multiple antigens in the same tissue and label antigens in tissues that had been stored for many years in epoxy resin. The technique allows quantitation of the gold label over plasma cell organelles and therefore gives information about the immunoglobulin secretory pathway in these cells. We found that the protein A-gold procedure compares favorably in technical ease with the immunoperoxidase, avidin-biotin peroxidase, and immunoglobulin-colloidal gold immunolabeling methods, and has added advantages in allowing precise localization and quantitation of the labeled antigen.
以往的研究发现，用常规方法制备的用于电子显微镜观察的组织中，免疫球蛋白无法进行免疫标记。为了验证这一结论，我们对用戊二醛固定、四氧化锇后固定并包埋于环氧树脂中的组织，采用了蛋白A-金后包埋免疫标记法；切片先用偏高碘酸钠预处理。我们还采用了多种常见的固定方案，并确定了免疫标记的最适宜条件。这项技术能够在人扁桃体、淋巴结、浆细胞瘤及肾活检组织中保存最佳且反差良好的浆细胞内对免疫球蛋白轻链进行超微结构定位。我们能够在同一组织中显示多种抗原，并对保存在环氧树脂中多年的组织中的抗原进行标记。该技术能够对浆细胞细胞器上的金标记进行定量，从而提供有关这些细胞中免疫球蛋白分泌途径的信息。我们发现，蛋白A-金法在操作简便性上与免疫过氧化物酶法、抗生物素蛋白-生物素过氧化物酶法及免疫球蛋白-胶体金免疫标记法相比毫不逊色，并且在能够对标记抗原进行精确定位和定量方面具有额外的优势。
