电子显微镜水平非同位素原位杂交技术的现状与问题

Non—Radioactive in situ Hybridization at Electron Microscopic Level: History,Current Situation and Problems

本文介绍原位杂交(非同位素方法)这一由分子生物学与组织学技术结合而成的新技术在电子显微镜水平的发展历史、目前的研究状况和存在的问题。本文结合作者应用电镜原位杂交技术探测肾脏内胶原mRNA分子的工作,报道了自己对包埋后电镜原位杂交技术的改良所作的探索,并阐述了对这一技术的灵敏度、"嗓音"干扰和分辨率等所存在问题的见解。

Non-radioactive in situ hybridization (ISH) has been extensively developed at light microscopic level during the past decade and currently becomes a routine technique. This technique at electron microscopic (EM) level, however, remained difficult,especially when aiming to detect non-viral mRNA. The history, current situation and problems of non-radioactive EM-ISH were reviewed in this article. The troubles involved in this teehique were sensitivity,noise interference,structural resolution,etc. The author's work on non-radioactive post-embedding EM-ISH was reported. The location of collagen type Ⅰ mRNA was determined on Lowieryl-K4M embedded kidney tissue using biotinylated or digoxidenin-laheled cDNA probe. The results were discussed in regard of enhancing signal intensity, discriminating non-specific labeling and improving structural resolution.