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In Vitro Detection and Quantification of Botulinum Neurotoxin Type E Activity in Avian Blood

机译：禽血中E型肉毒杆菌神经毒素的体外检测和定量

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摘要

Botulinum neurotoxin serotype E (BoNT/E) outbreaks in the Great Lakes region cause large annual avian mortality events, with an estimated 17,000 bird deaths reported in 2007 alone. During an outbreak investigation, blood collected from bird carcasses is tested for the presence of BoNT/E using the mouse lethality assay. While sensitive, this method is labor-intensive and low throughput and can take up to 7 days to complete. We developed a rapid and sensitive in vitro assay, the BoTest Matrix E assay, that combines immunoprecipitation with high-affinity endopeptidase activity detection by F?rster resonance energy transfer (FRET) to rapidly quantify BoNT/E activity in avian blood with detection limits comparable to those of the mouse lethality assay. On the basis of the analysis of archived blood samples ( n = 87) collected from bird carcasses during avian mortality investigations, BoTest Matrix E detected picomolar quantities of BoNT/E following a 2-h incubation and femtomolar quantities of BoNT/E following extended incubation (24 h) with 100% diagnostic specificity and 91% diagnostic sensitivity.
机译：大湖地区爆发的E型肉毒神经毒素血清型（BoNT / E）导致每年发生大量禽类死亡事件，仅2007年一年就报告死亡禽鸟17,000人。在暴发调查期间，使用小鼠致死率检测从禽尸体收集的血液中是否存在BoNT / E。这种方法虽然很敏感，但劳动强度大且通量低，最多可能需要7天才能完成。我们开发了一种快速灵敏的体外测定法，即BoTest Matrix E测定法，该方法将免疫沉淀与高亲和力内肽酶活性检测（通过Fster共振能量转移（FRET））相结合，可快速定量禽血中BoNT / E活性，且检测限相当与小鼠致死率测定的结果相同。根据在禽类死亡调查期间从鸟类尸体收集的存档血液样本（n = 87）的分析，BoTest Matrix E在2 h孵育后检测到了皮摩尔量的BoNT / E，在长时间孵育后检测到了飞摩尔量的BoNT / E。 （24 h）具有100％的诊断特异性和91％的诊断敏感性。

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