一种色光蛋白,具有很大的斯托克斯转移,用于单激发多色FCCS和FRET成像
Daria M Shcherbakova1, Mark A Hink, Linda Joosen
1Department of Anatomy and Structural Biology and Gruss-Lipper Biophotonics Center, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, New York 10461, USA.
Journal of the American Chemical Society
|April 11, 2012
概括
研究人员开发了LSSmOrange,这是一种明亮的色光蛋白 (FP),可以填补光谱间隙. 这项创新能够在活细胞中实现先进的多色成像和单分子分析.
科学领域:
- 细胞和分子成像技术
- 生物物理学的生物物理.
- 蛋白质工程是指蛋白质工程.
背景情况:
- 基因编码的光蛋白 (FPs) 是活细胞动态过程研究的重要工具.
- 现有的FP调色板具有光谱差距,限制了多色成像能力.
研究的目的:
- 引入一种具有较大的斯托克斯转移 (LSS) 的新型单体色FP,命名为LSSmOrange.
- 为了证明LSSmOrange在先进的多色成像和单分子分析应用中的实用性.
主要方法:
- LSSmOrange的发展和特征 (在437/572 nm的激发/发射).
- 在四色单激光光交叉相关谱学 (FCCS) 中的应用.
- 用于使用CFP-YFP和LSSmOrange-mKate2生物传感器进行福斯特共振能量转移 (FRET) 成像.
主要成果:
- LSSmOrange的亮度相当于绿黄色FP的亮度,明显高于红色LSSFP的亮度.
- 启用了新的四色FCCS用于定量单分子分析.
- 使用单一激发波长为两个FRET对 (包括CFP-YFP) 提供了同时成像的便利.
- 实时成像证明了亡活动和的波动.
结论:
- LSSmOrange有效地填补了绿黄色和红色LSSFP之间的光谱差距.
- 它的亮度和光谱特性使新的多色应用和先进的生物物理技术成为可能.
- 研究揭示了激发状态质子转移机制,该机制负责LSS表型.
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