Terasaki M, Reese T S
Laboratory of Neurobiology, NINDS, NIH, Betheda, MD 20892.
J Cell Sci. 1992 Feb;101 ( Pt 2):315-22. doi: 10.1242/jcs.101.2.315.
The original concept of endoplasmic reticulum derived from the observation of a reticular network in cultured fibroblasts by electron microscopy of whole cells. It was previously reported that the fluorescent dye, DiOC6(3), stains a similar network as well as mitochondria and other organelles in living cells. Here, we investigate the significance of the structures labeled by DiO6(3) in CV-1 cells, a monkey epithelial cell line. First, we show that the network stained in living CV-1 cells is preserved by glutaraldehyde fixation and then we co-label it with an antibody against BiP (immunoglobulin binding protein), a protein commonly accepted to be present in the endoplasmic reticulum. Anti-BiP labeled the same network as that labeled by DiOC6(3), so this network now is identified as being part of the endoplasmic reticulum. DiOC6(3) labels many other membrane compartments in addition to the endoplasmic reticulum. This, along with its lipophilic properties, suggests that DiOC6(3) stains all intracellular membranes. However, the extensive reticular network in the thin peripheral regions of cultured cells is easily distinguished from these other membranes. Thus, staining by DiOC6(3) is a useful method for localizing the endoplasmic reticulum, particularly in thin peripheral regions of cultured cells.
内质网的最初概念源自对培养的成纤维细胞进行全细胞电子显微镜观察时所发现的网状网络。此前有报道称，荧光染料DiOC6(3)可对活细胞中的类似网络以及线粒体和其他细胞器进行染色。在此，我们研究了DiO6(3)在猴上皮细胞系CV-1细胞中标记的结构的意义。首先，我们发现戊二醛固定可保留活CV-1细胞中染色的网络，然后我们用抗BiP（免疫球蛋白结合蛋白）抗体对其进行共标记，BiP是一种普遍认为存在于内质网中的蛋白质。抗BiP标记的网络与DiOC6(3)标记的网络相同，因此该网络现在被确定为内质网的一部分。除了内质网，DiOC6(3)还标记许多其他膜区室。这一点连同其亲脂性表明，DiOC6(3)可对所有细胞内膜进行染色。然而，培养细胞薄周边区域中的广泛网状网络很容易与这些其他膜区分开来。因此，用DiOC6(3)染色是一种定位内质网的有用方法，尤其是在培养细胞的薄周边区域。
