膜电喷雾质谱法快速检测临床样本中志贺毒素

Rapid determination of shiga toxin in clinical samples using membrane electrospray ionization mass spectrometry

志贺毒素是大肠杆菌O157:H7重要的毒力因子,是毒力最强的毒素之一。本文建立了采用膜电喷雾质谱(MESI-MS/MS)快速检测人粪便样本中志贺毒素的方法。将两种志贺毒素的特异性肽段作为检测目标物,在蛋白水平上对志贺毒素进行检测。对肽段YNDDDTFTVK(YK1)和YNEDDTFTVK(YK2)的母子离子对建立了定量检测方法,并对检测方法学进行了验证。实验结果表明方法具有良好的检测精密度、准确度以及回收率。MESI-MS/MS具有去除基质效应的能力,可对酶解后的混合蛋白样本实现直接检测,全部操作过程可在3 h内完成。方法有望推广至其它细菌毒素的检测,且有望与便携式质谱相结合实现现场快速检测。

Shiga toxin is an important virulence factor of Escherichia coli O157： H7,and it is one of the most virulent toxins.Developing rapid detection methods of Shiga toxin are essential requirement for public health and clinical testing.In this study,a quantitative method was developed to determine Shiga toxin based on their unique peptides using MESI-MS/MS.Two unique peptides Stx1 and Stx2 were used as the detection targets,and the Stx is able to be detected on the protein level.Precursor/product-ion pairs of peptide YNDDDTFTVK（YK1） and YNEDDTFTVK（YK2） have been selected,and the methodology has been validated.The experimental results showed that the method had good precision,accuracy and recovery.MESI-MS/MS has the ability to remove matrix effects.As a result,the crude sample after trysin digestion can be analyzed directly,and the whole process can be completed within 3 hours.This simple and rapid detection method is expected to be extended to other bacterial toxins detection,and can be combined with portable mass spectrometry to achieve on-site rapid detection.