Kahana J A, Silver P A
Dana-Farber Cancer Institute, Boston, Massachusetts, USA.
Curr Protoc Pharmacol. 2001 May;Chapter 6:Unit 6.6. doi: 10.1002/0471141755.ph0606s05.
Fluorescent molecules serve as valuable tools for the detection of a variety of biochemical phenomena. Such reagents have been employed for protein localization, quantitation of gene expression, detection of nucleic acids, cell sorting, and determination of chemical concentrations. Although fluorescence is a useful tool for detecting molecules within cells, its application in vivo has been limited. The ideal vital fluorescent tag should (1) be detectable without causing cytological damage, (2) be able to label a wide variety of cell types readily, and (3) be able to be targeted to virtually any subcellular region. The recently cloned green fluorescent protein (GFP) from the jellyfish Aequorea victoria is such a molecule. This overview describes the use of this proteinaceous fluorophore for in vivo observation of cellular phenomena, including applications and problems with the use of GFP, a discussion of mutant GFPs with altered fluorescence characteristics, and also some details on microscopy requirements.
荧光分子是检测各种生化现象的宝贵工具。此类试剂已用于蛋白质定位、基因表达定量、核酸检测、细胞分选以及化学浓度测定。尽管荧光是检测细胞内分子的有用工具，但其在体内的应用一直受到限制。理想的活体荧光标记物应具备以下条件：（1）可检测且不造成细胞损伤；（2）能够轻松标记多种细胞类型；（3）能够靶向几乎任何亚细胞区域。最近从维多利亚多管水母中克隆出的绿色荧光蛋白（GFP）就是这样一种分子。本综述描述了这种蛋白质荧光团在体内观察细胞现象中的应用，包括GFP的使用方法和问题、对具有改变荧光特性的突变型GFP的讨论，以及一些关于显微镜要求的细节。
