A Rapid, Sensitive and Fully Automated Anthrax Test

描述(申请人提供)：生物恐怖主义和生物武器是对我们国家的威胁。这种威胁在2001年的炭疽热袭击中变成了现实。虽然袭击规模不大，但其影响达到了恐怖的程度。不幸的是，正如前参议员沃伦·鲁德曼所指出的，“未来几年在这个国家发生恐怖袭击的可能性很大，而不是不可能”。美国外交关系委员会最近的研究得出结论，我们的国家仍然“没有准备好应对美国领土上的灾难性袭击，特别是涉及化学、生物或核毒剂的袭击”。 在发生生物恐怖主义袭击时，最重要的第一反应之一是快速识别一个或多个生物制剂，并及时诊断那些被感染者。目前用于炭疽杆菌敏感检测的方法是聚合酶链式反应(PCR)方法。然后用培养方法和/或EIA确认该有机体。然而，聚合酶链式反应的使用也并非没有问题。聚合酶链式反应扩增产物可能受到污染，这就需要一个专门的、高度受控的中心实验室，而这又需要长途运送样品。事实上，聚合酶链式反应容易受到抑制性杂质的影响，这需要长时间的样品准备和重复测试。所有这些问题都是在2001年炭疽热恐怖袭击期间遇到的。 我们已经开发出一种快速(大约60分钟)和敏感(不到1个细菌)的败血症检测原型，使用我们的基于微粒的扩增(MBA)技术和一类新的细菌标志物。这一阶段应用的具体目的是研究开发炭疽芽孢杆菌检测系统的可行性，该系统包括用于检测营养细胞的核酸(pXO1和pXO2质粒)检测和用于检测炭疽芽胞的凝集试验。这两种检测方法使用相同的MBA技术，并且具有(A)快速(60-90分钟)、(B)灵敏(核酸检测为100cU，凝集检测为5000个孢子)和(C)特异性(几乎没有交叉反应)。

DESCRIPTION (provided by applicant): Bioterrorism and biowarfare are a threat for our nation. Such threat became reality in the 2001 Anthrax attack. Although the attack occurred at a small scale, its impact reached terror proportion. Unfortunately, as former Senator Warren Rudman pointed out, "the likelihood of a terrorist attack in this country in the next several years is more likely than unlikely". Recent study by the Council on Foreign Relations concluded that our nation remains "unprepared to handle a catastrophic attack on American soil, particularly one involving chemical, biological or nuclear agents". In the event of a Bioterrorism attack, one of the most important first responses is rapid identification of the biological agent or agents, and timely diagnosis of those who have been infected. Current methods of choice for sensitive detection of B. anthracis are those involving polymerase chain reaction (PCR). The organism is then confirmed with culture methods and/or EIA. The use of PCR is not without problems, however. Possible contamination of PCR amplicons necessitates a specialized and highly controlled central laboratory, which in turn necessitates long distance sample shipping. The fact that PCR is susceptible to inhibitory impurities requires lengthy sample preparation and duplicate testing. All these problems had been encountered during the 2001 Anthrax terror attack. We have developed a rapid (approximately 60 minutes) and sensitive (less than 1 bacterium) prototype sepsis test using our microparticle based amplification (MBA) technology and a novel class of bacterial markers. The specific aims for this Phase I application is to study the feasibility of developing a Bacillus anthracis test system comprising a nucleic acids (pXO1 and pXO2 plasmids) test for the detection of vegetative cells and an agglutination assay for the detection of B. anthracis spores. Both assays use the same MBA technology and are (a) rapid (60-90 minutes), (b) sensitive (< 100 CPU for the nucleic acid test and <5000 spores for the agglutination assay), and (c) specific (little cross reactivity).