Microfluidic Mixers for the Investigation of Rapid Protein Folding Kinetics Using Synchrotron Radiation Circular Dichroism Spectroscopy

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• 作者：Avinash S. Kane ; Armin Hoffmann ; Peter Baumgrtel ; Robert Seckler ; Gerd Reichardt ; David A. Horsley ; Benjamin Schuler ; Olgica Bakajin
• 刊名：Analytical Chemistry
• 出版年：2008
• 出版时间：December 15, 2008
• 年：2008
• 卷：80
• 期：24
• 页码：9534-9541
• 全文大小：300K
• 年卷期：v.80,no.24(December 15, 2008)
• ISSN：1520-6882

文摘

We have developed a microfluidic mixer optimized for rapid measurements of protein folding kinetics using synchrotron radiation circular dichroism (SRCD) spectroscopy. The combination of fabrication in fused silica and synchrotron radiation allows measurements at wavelengths below 220 nm, the typical limit of commercial instrumentation. At these wavelengths, the discrimination between the different types of protein secondary structure increases sharply. The device was optimized for rapid mixing at moderate sample consumption by employing a serpentine channel design, resulting in a dead time of less than 200 μs. Here, we discuss the design and fabrication of the mixer and quantify the mixing efficiency using wide-field and confocal epi-fluorescence microscopy. We demonstrate the performance of the device in SRCD measurements of the folding kinetics of cytochrome c, a small, fast-folding protein. Our results show that the combination of SRCD with microfluidic mixing opens new possibilities for investigating rapid conformational changes in biological macromolecules that have previously been inaccessible.