National Center for Environmental Health, Centers for Disease Control and Prevention, Atlanta, GA 30341, USA.
Anal Biochem. 2011 May 1;412(1):67-73. doi: 10.1016/j.ab.2011.01.025. Epub 2011 Jan 26.
Botulinum neurotoxins (BoNTs) are the most toxic substances known to humankind. Rapid and sensitive detection of BoNTs is necessary for timely clinical confirmation of the disease state in botulism. BoNTs cleave proteins and peptide mimics at specific sites. A mass spectrometry (MS)-based method, Endopep-MS, can detect these cleavages and has detection limits of 0.05-0.5 mouse LD(50) (U) in serum, depending on the BoNT serotypes. In this method, the products generated from cleavage of peptide substrates using antibody affinity-purified toxins are detected by MS. Nonspecific bound endogenous proteases or peptidases in stool can coextract with the toxin, cleaving the peptide substrates and reducing the sensitivity of the method. Here we report a method to reduce nonspecific substrate cleavage by reducing stool protease coextraction in the Endopep-MS assay.
肉毒神经毒素（BoNTs）是人类已知的最毒物质。快速、灵敏地检测 BoNTs 对于及时确认肉毒中毒患者的疾病状态非常必要。BoNTs 可以在特定位置切割蛋白质和肽模拟物。基于质谱（MS）的方法 Endopep-MS 可以检测这些切割，并且根据 BoNT 血清型的不同，在血清中的检测限为 0.05-0.5 小鼠 LD(50)（U）。在该方法中，使用抗体亲和纯化毒素切割肽底物产生的产物通过 MS 检测。粪便中与毒素一起共提取的非特异性结合的内源性蛋白酶或肽酶会切割肽底物，从而降低该方法的灵敏度。在这里，我们报告了一种通过减少 Endopep-MS 检测中粪便蛋白酶共提取来减少非特异性底物切割的方法。
