Scarlata Suzanne, Dowal Louisa
Department of Physiology and Biophysics, State University of New York at Stony Brook, USA.
Methods Mol Biol. 2004;237:223-32. doi: 10.1385/1-59259-430-1:223.
A major advance in biology is the ability to attach either green fluorescence protein (GFP) or one of its variants to a target protein and follow its cellular localization and interaction with other partners by fluorescence microscopy. Our laboratory has previously developed fluorescence energy-transfer methods to measure the kinetics and affinities of the lateral association between phospholipase C (PLC) and G protein subunits on membrane surfaces. We are currently developing methods to view these associations in living cells using fluorescence resonance energy transfer (FRET) between GFP-based chimeras. Because the improvements and variations of these GFP-based FRET techniques has continued on a rapid pace, we focus only on the basic principles behind these measurements and the methods used, which may continue to be applicable as improvements become available.
生物学领域的一项重大进展是能够将绿色荧光蛋白（GFP）或其变体之一附着到目标蛋白上，并通过荧光显微镜追踪其细胞定位以及与其他蛋白的相互作用。我们实验室之前开发了荧光能量转移方法，以测量磷脂酶C（PLC）与膜表面G蛋白亚基之间横向结合的动力学和亲和力。我们目前正在开发利用基于GFP的嵌合体之间的荧光共振能量转移（FRET）在活细胞中观察这些结合的方法。由于这些基于GFP的FRET技术的改进和变体一直在快速发展，我们仅关注这些测量背后的基本原理以及所使用的方法，随着技术的不断改进，这些原理和方法可能仍然适用。
