The Graduate School for the Creation of New Photonics Industries, 1955-1 Kurematsu-cho, Nishi-ku, Hamamatsu, Shizuoka 431-1202, Japan.
Hamamatsu Photonics K.K., 314-5 Shimokanzo, Iwata, Shizuoka 438-0193, Japan.
J Am Soc Mass Spectrom. 2020 Oct 7;31(10):2154-2160. doi: 10.1021/jasms.0c00255. Epub 2020 Sep 2.
DIUTHAME (desorption ionization using through hole alumina membrane) is a novel matrix-free laser desorption/ionization method that enables highly reproducible acquisition of mass spectra. This study aims to evaluate the applicability of DIUTHAME to the acetylcholinesterase reaction assay (AChE assay) commonly used in high-throughput screening (HTS) in the drug discovery process. The commercially available 9-ch DIUTHAME chip was applied to a series of AChE assays prepared with different reaction times. Numerous positive-mode TOF mass spectra were acquired from the raster-scanned sample spots of the AChE assays to analyze the progress of the enzyme reaction and to perform mass spectrometry imaging of the sample spots. The reaction kinetics plots obtained by DIUTHAME were found to reflect the time course of reaction progression as much as those obtained by MALDI and were found to have less error variation than MALDI. DIUTHAME allowed the already irradiated sample spots to be reused to reproduce the reaction kinetics plots by the second measurement conducted a week after, whereas MALDI was not able to properly reproduce the kinetics plots by remeasuring the irradiated sample spots. In DIUTHAME, which does not require a matrix, the experimental procedure for measuring AChE assay becomes extremely simple. DIUTHAME is potentially able to provide more precise AChE reaction kinetics plots than what have been accomplished by MALDI.
DIUTHAME（通过通孔氧化铝膜解吸电离）是一种新颖的无基质激光解吸/电离方法，可实现高度重现的质谱采集。本研究旨在评估 DIUTHAME 在药物发现过程中高通量筛选（HTS）中常用的乙酰胆碱酯酶反应测定（AChE 测定）中的适用性。商用的 9-ch DIUTHAME 芯片应用于一系列具有不同反应时间的 AChE 测定中。从 AChE 测定的光栅扫描样品点中获得了大量正离子模式 TOF 质谱，以分析酶反应的进展，并对样品点进行质谱成像。通过 DIUTHAME 获得的反应动力学图与 MALDI 获得的动力学图一样，能够很好地反映反应进展的时间过程，并且比 MALDI 具有更小的误差变化。DIUTHAME 允许已经辐照的样品点被重复使用，通过第二周的第二次测量来重现反应动力学图，而 MALDI 无法通过重新测量辐照样品点来正确重现动力学图。在不需要基质的 DIUTHAME 中，测量 AChE 测定的实验程序变得极其简单。DIUTHAME 有可能提供比 MALDI 更精确的 AChE 反应动力学图。
