Detection of functional ricin by immunoaffinity and liquid chromatography-tandem mass spectrometry
Detection of functional ricin by immunoaffinity and liquid chromatography-tandem mass spectrometry
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DOI：
10.1021/ac061498b
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发表时间：
2007-01-15
影响因子：
7.4
通讯作者：
Ezan, E.
中科院分区：
文献类型：
--
作者：
Becher, F.;Duriez, E.;Ezan, E.
蓖麻毒素是一种可能用于生物恐怖目的的生物武器。蓖麻毒素是核糖体失活蛋白(RIPs)家族中的一员，其A链具有N-糖苷酶活性，不可逆地抑制蛋白质合成。在本文中，我们证明，只要使用适当的样品制备，这种酶活性可以用于功能性蓖麻毒素的检测，其灵敏度和特异性类似于最佳的ELISA法，并允许应用于环境样品。蓖麻毒素首先被针对B链的单抗捕获，然后固定在磁珠上。然后，通过使用液相色谱与串联质谱联用来确定由A链从RNA模板中释放的腺嘌呤来实现检测。免疫亲和步骤与酶活性检测相结合，可以对整个功能性蓖麻毒素进行特异性检测，在500亩L样品中浓缩毒素后，检测下限为0.1 ng/mL(1.56 pm)。该方法的变异系数为10%。最后，将该方法成功地应用于牛奶和自来水或瓶装水样中。
The toxin ricin is a biological weapon that may be used for bioterrorist purposes. As a member of the group of ribosome-inactivating proteins (RIPs), ricin has an A-chain possessing N-glycosidase activity which irreversibly inhibits protein synthesis. In this paper, we demonstrate that provided appropriate sample preparation is used, this enzymatic activity can be exploited for functional ricin detection with sensitivity similar to the best ELISA and specificity allowing application to environmental samples. Ricin is first captured by a monoclonal antibody directed against the B chain and immobilized on magnetic beads. Detection is then realized by determination of the adenine released by the A chain from an RNA template using liquid chromatography coupled to tandem mass spectrometry. The immunoaffinity step combined with the enzymatic activity detection leads to a specific assay for the entire functional ricin with a lower limit of detection of 0.1 ng/mL (1.56 pM) after concentration of the toxin from a 500 mu L sample size. The variability of the assay was 10%. Finally, the method was applied successfully to milk and tap or bottled water samples.