引用

炭疽芽孢杆菌是一种主要的生物战威胁。吸入性感染可以迅速致死。虽然抗生素治疗是有效的，但如果延误诊断，迅速产生的毒素可能已经达到致死量。本文介绍了一种快速、灵敏、特异、准确的检测血浆中炭疽芽孢杆菌活动性感染的方法。其中一个毒力因子，炭疽致死因子，是一种存在于感染早期血液中的内肽酶。然而，在血浆中使用肽底物检测内肽酶是有问题的，因为存在其他蛋白酶和底物的非特异性裂解的可能性。本研究设计的荧光标记肽底物MAPKKide Plus不被血浆蛋白酶裂解，对致死因子具有特异性。描述了三种检测策略。两种方法包括使用致命因子抗体包被微量滴度板或类似的包被免疫管从血浆中捕获富集。将捕获的致死因子暴露在MAPKKide Plus中，并通过HPLC或微孔板阅读器确定裂解量。 Concentration of lethal factor using the antibody-coated plates aplnd HPLC allows for detection of less than 5 pg lethal factor/ml of neat plasma after 2 hours of incubation. Using antibody-coated immuno-tubes, 20 pg lethal factor/ml plasma can be detected in 5 hours by a simple end point read of fluorescence in a microplate reader. For a third strategy, the substrate is added directly to diluted plasma, and cleavage is monitored by the increase in fluorescence as a function of time. The limit of detection by this simple method is 25 ng lethal factor/ml of plasma in 15 minutes, 5 ng/ml after 45 minutes, and <1 ng lethal factor/ml of plasma after 5 hours.