Hertzog David E, Michalet Xavier, Jäger Marcus, Kong Xiangxu, Santiago Juan G, Weiss Shimon, Bakajin Olgica
Mechanical Engineering Department, Stanford University, Stanford, California 94305, USA.
Anal Chem. 2004 Dec 15;76(24):7169-78. doi: 10.1021/ac048661s.
We have developed a microfluidic mixer for studying protein folding and other reactions with a mixing time of 8 mus and sample consumption of femtomoles. This device enables us to access conformational changes under conditions far from equilibrium and at previously inaccessible time scales. In this paper, we discuss the design and optimization of the mixer using modeling of convective diffusion phenomena and a characterization of the mixer performance using microparticle image velocimetry, dye quenching, and Forster resonance energy-transfer (FRET) measurements of single-stranded DNA. We also demonstrate the feasibility of measuring fast protein folding kinetics using FRET with acyl-CoA binding protein.
我们开发了一种微流体混合器,用于研究蛋白质折叠和其他反应,其混合时间为8微秒,样品消耗量为飞摩尔。该装置使我们能够在远离平衡的条件下以及在以前无法达到的时间尺度上观察构象变化。在本文中,我们讨论了使用对流扩散现象建模对混合器进行的设计和优化,以及使用微粒图像测速技术、染料猝灭和单链DNA的荧光共振能量转移(FRET)测量对混合器性能进行的表征。我们还展示了使用FRET结合酰基辅酶A结合蛋白测量快速蛋白质折叠动力学的可行性。