Veltel D, Robenek H
Cell Biology Group, Faculty of Medicine, University of Münster, FRG.
J Histochem Cytochem. 1988 Oct;36(10):1295-303. doi: 10.1177/36.10.3418108.
We studied the distribution of acetylcholine receptors (AchRs) in rat skeletal muscles, Torpedo membrane fractions, and cultured rat skeletal myotubes. AchRs were first exposed to alpha-bungarotoxin (alpha-BTX) followed by anti-alpha-BTX antibodies. Bound antibodies were visualized with FITC-conjugated goat anti-rabbit antibodies for light microscopy or with peroxidase-conjugated goat anti-rabbit or gold-labeled protein A for electron microscopy. The protocol developed for the present studies detected AchRs with high specificity. In addition, we combined post-fixation immunogold cytochemical and surface replication techniques to study the distribution of AchRs on the dorsal surface of cultured myotubes in detail. Two distinct distribution patterns of AchRs on the cell surfaces of the myotubes were revealed; AchRs were either diffusely distributed or clustered. Dispersed AchRs usually surrounded clusters of AchRs. The AchRs in the clusters were characteristically arranged, and small aggregates of AchRs could also be observed within the clusters. The techniques used in this study are appropriate for studying the dynamics of AchR clustering.
我们研究了乙酰胆碱受体（AchRs）在大鼠骨骼肌、电鳐膜组分以及培养的大鼠骨骼肌肌管中的分布情况。首先让AchRs与α-银环蛇毒素（α-BTX）接触，然后再与抗α-BTX抗体接触。结合的抗体通过异硫氰酸荧光素（FITC）偶联的山羊抗兔抗体进行可视化以便于光学显微镜观察，或者通过过氧化物酶偶联的山羊抗兔抗体或金标记的蛋白A进行可视化以便于电子显微镜观察。为本研究开发的方案能够以高特异性检测AchRs。此外，我们结合了后固定免疫金细胞化学和表面复型技术，以详细研究AchRs在培养肌管背表面的分布。在肌管的细胞表面揭示了AchRs的两种不同分布模式；AchRs要么是分散分布，要么是聚集分布。分散的AchRs通常围绕着AchRs簇。簇中的AchRs具有特征性排列，并且在簇内也可以观察到AchRs的小聚集体。本研究中使用的技术适用于研究AchR聚集的动态过程。
