Currently, all assays measuring acetylcholinesterase (AChE) activity following a suspected nerve agent exposure leverage methodologies that fail to identify the agent. This limits the overall effectiveness and ability to administer proper countermeasures. As such, there is an urgent need to identify novel, rapid, and more comprehensive approaches to establish AChE activity, including identification of the toxicant. Paper spray mass spectrometry was used to monitor the activity of acetylcholinesterase, both in-solution and on modified hydrophobic paper surface. Hydrophobic paper surfaces were prepared using vaporized trichloro(3,3,3-trifluoropropyl)silane. In both approaches, mixtures of diluted human whole blood with and without VX were mixed with a non-endogenous AChE specific substrate, 1,1-dimethyl-4-acetylthiomethylpiperidinium (MATP+). Formation of the cleaved MATP+ product was monitored over time and compared to MATP+ to determine relative AChE activity. This on-substrate assay was effective at determining AChE activity and identifying the toxicant; however, determination of AChE activity in-solution proceeded at a slower rate. The on-substrate assay serves as a pioneering example of an enzymatic reaction occurring on the surface of a paper spray ionization ticket. This work broadens the range of applications relating to paper spray ionization-based clinical diagnostic assays.

Open image in new window

目前，在疑似神经毒剂暴露后测量乙酰胆碱酯酶 (AChE) 活性的所有测定均利用无法识别该毒剂的方法。这限制了采取适当对策的整体有效性和能力。因此，迫切需要确定新颖、快速和更全面的方法来确定 AChE 活性，包括毒物的鉴定。使用纸喷雾质谱法来监测溶液中和改性疏水纸表面上的乙酰胆碱酯酶的活性。使用汽化的三氯(3,3,3-三氟丙基)硅烷制备疏水纸表面。在这两种方法中，将含有和不含 VX 的稀释人全血混合物与非内源性 AChE 特异性底物 1,1-二甲基-4-乙酰硫基甲基哌啶 (MATP+) 混合。随着时间的推移，监测切割的 MATP+ 产物的形成，并与 MATP+ 进行比较，以确定相对 AChE 活性。这种底物检测可有效测定 AChE 活性并识别毒物；然而，溶液中 AChE 活性的测定速度较慢。基质上测定是纸喷雾电离票表面发生酶反应的开创性示例。这项工作拓宽了与基于纸喷雾电离的临床诊断测定相关的应用范围。

在新窗口中打开图像