Center for Hybrid Nanostructures (CHyN), Universität Hamburg, 22761 Hamburg, Germany.
HAMBURG SCHOOL OF FOOD SCIENCE - Institute of Food Chemistry, Universität Hamburg, 20146 Hamburg, Germany.
Phys Chem Chem Phys. 2023 Mar 8;25(10):7312-7322. doi: 10.1039/d2cp02361j.
Top-down proteomics deals with the characterization of intact biomolecules, which reduces the sample complexity and facilitates the detection of modifications at the protein level. The combination of the matrix-assisted laser desorption/ionization (MALDI) technique with time-of-flight (TOF) mass analysis allows for the generation of gaseous ions in low charge states from high-mass biomolecules, followed by their mass-to-charge ratio (/) separation, as high-mass ions drift down the flight tube more slowly than lighter ones. However, the detection efficiency of conventional microchannel plate (MCP) detectors is strongly reduced with decreasing ion velocity-corresponding to an increase in ion mass-which impedes the reliable detection of high-mass biomolecules. Herein, we present a simple modification of the MCP detector that allows for the amplification of the signal from ionized proteins of up to / 150 000. Two circular electrodes were assembled in front of the conventional detector and set to negative electrical voltages to affect the positively charged ions directly before they impinge on the MCP, possibly through a combination of a velocity boost and ion optical effects. In the present study, three booster electrode configurations were experimentally tested to maximize the signal intensification. Compared to the conventional MCP assembly, the signal intensity was amplified in a proof-of-concept experiment by a factor of 24.3 and of 10.7 for the singly charged BSA ion (/ 66 400) and for the singly charged IgG ion (/ 150 000), respectively, by applying the booster-MCP (BMCP) detector.
自上而下的蛋白质组学涉及完整生物分子的特征描述,这降低了样品的复杂性,并有利于在蛋白质水平检测修饰。基质辅助激光解吸/电离(MALDI)技术与飞行时间(TOF)质谱分析的结合允许从高质量的生物分子中产生低电荷状态的气态离子,然后对其进行质量电荷比(/)分离,因为高质量的离子在飞行管中下降的速度比轻离子慢。然而,传统微通道板(MCP)检测器的检测效率随着离子速度的降低而大大降低-这对应于离子质量的增加-从而阻碍了对高质量生物分子的可靠检测。在此,我们对 MCP 检测器进行了简单的改进,使离子化蛋白质的信号放大高达/ 150 000。两个圆形电极组装在传统检测器的前面,并设置为负的电电压,以直接影响在 MCP 撞击之前带正电荷的离子,可能通过速度提升和离子光学效应的组合。在本研究中,实验测试了三种升压电极配置,以最大限度地提高信号增强。与传统的 MCP 组件相比,通过应用升压-MCP(BMCP)检测器,在概念验证实验中,BSA 离子(/ 66400)的单电荷离子和 IgG 离子(/ 150000)的单电荷离子的信号强度分别放大了 24.3 倍和 10.7 倍。