An integrated laser-tweezers instrument for microanalysis of individual protein aggregates
An integrated laser-tweezers instrument for microanalysis of individual protein aggregates
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DOI：
10.1016/j.snb.2007.09.052
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发表时间：
2008-02-22
影响因子：
8.4
通讯作者：
Luchette, Paul
中科院分区：
文献类型：
--
作者：
Mao, Hanbin;Luchette, Paul
在此，我们介绍了一种新的激光镊子仪器，能够提供单独捕获的蛋白质聚集体的物理和化学信息。通过研究直径为纳米到微米的α -弹性蛋白液滴的聚集过程，证明了这些能力。在1064nm近红外（NIR）激光聚焦下，该仪器捕获了聚集弹性蛋白分子的单个液滴。这些单个弹性蛋白聚集体的大小是通过使用位置敏感光电探测器（PSPD）记录它们在100 kHz频率下的热运动来推断的。利用532 nm激光激发的共聚焦荧光和共聚焦拉曼光谱揭示了捕获粒子的化学信息。被捕获粒子外的背景信号通过后反射几何中的针孔消除。物理和共聚焦荧光测量显示，在单个弹性蛋白聚集体中，快速过程的转变温度为38摄氏度，这一值与团块的云温度一致。共聚焦拉曼光谱结果表明，弹性蛋白水两相体系（ATPS）形成过程缓慢，转变温度为35℃。(C) 2007 Elsevier B.V.版权所有
Herein we introduce a new laser-tweezers instrument capable of providing both physical and chemical information of individually trapped protein aggregates. These capabilities were demonstrated by investigating the aggregation process of alpha-elastin droplets of nanometers to micrometers in diameter. Individual droplets of aggregated elastin molecules were trapped at the focus of a 1064 nm near infrared (NIR) laser in the instrument. The size of these individual elastin aggregates was inferred by recording their thermal motion at a frequency of 100 kHz using a position sensitive photodetector (PSPD). The chemical information of the trapped particles was revealed by confocal fluorescence and confocal Raman spectroscopies using 532-nm laser excitation. Background signals outside trapped particles were eliminated by a pinhole in back reflected geometry. Both physical and confocal fluorescence measurements revealed a transition temperature of 38 degrees C for a fast process in individual elastin aggregates, a value consistent with the clouding temperature in bulk. Confocal Raman spectroscopy yielded a transition temperature of 35 degrees C for a slow process characteristic of the formation of elastin aqueous two phase systems (ATPS). (C) 2007 Elsevier B.V. All rights reserved.