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首页> 美国卫生研究院文献>Clinical and Diagnostic Laboratory Immunology >Detection of Francisella tularensis in Biological Specimens Using a Capture Enzyme-Linked Immunosorbent Assay an Immunochromatographic Handheld Assay and a PCR
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Detection of Francisella tularensis in Biological Specimens Using a Capture Enzyme-Linked Immunosorbent Assay an Immunochromatographic Handheld Assay and a PCR

机译：使用捕获酶联免疫吸附测定免疫色谱手持测定和PCR检测生物标本中的弗拉杆菌

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The early detection of Francisella tularensis, the causative agent of tularemia, is important for adequate treatment by antibiotics and the outcome of the disease. Here we describe a new capture enzyme-linked immunosorbent assay (cELISA) based on monoclonal antibodies specific for lipopolysaccharide (LPS) of Francisella tularensis subsp. holarctica and Francisella tularensis subsp. tularensis. No cross-reactivity with Francisella tularensis subsp. novicida, Francisella philomiragia, and a panel of other possibly related bacteria, including Brucella spp., Yersinia spp., Escherichia coli, and Burkholderia spp., was observed. The detection limit of the assay was 103 to 104 bacteria/ml. This sensitivity was achieved by solubilization of the LPS prior to the cELISA. In addition, a novel immunochromatographic membrane-based handheld assay (HHA) and a PCR, targeting sequences of the 17-kDa protein (TUL4) gene of F. tularensis, were used in this study. Compared to the cELISA, the sensitivity of the HHA was about 100 times lower and that of the PCR was about 10 times higher. All three techniques were successfully applied to detect F. tularensis in tissue samples of European brown hares (Lepus europaeus). Whereas all infected samples were recognized by the cELISA, those with relatively low bacterial load were partially or not detected by PCR and HHA, probably due to inhibitors or lack of sensitivity. In conclusion, the HHA can be used as a very fast and simple approach to perform field diagnosis to obtain a first hint of an infection with F. tularensis, especially in emergent situations. In any suspect case, the diagnosis should be confirmed by more sensitive techniques, such as the cELISA and PCR.
机译：早期发现图拉弗朗西斯的病因是弗朗西斯菌Tularensis，对于通过抗生素进行充分治疗和疾病的结果非常重要。在这里，我们描述了一种新的捕获酶联免疫吸附测定法（cELISA），该测定法基于土拉弗朗西斯菌亚种脂多糖（LPS）特异的单克隆抗体。 holarctica和土拉弗朗西斯菌。吐拉ensis与土拉弗朗西斯菌无亚种的交叉反应。观察到新生病毒，费氏弗朗西斯菌和其他可能相关的细菌，包括布鲁氏菌属，耶尔森菌属，大肠杆菌和伯克霍尔德氏菌。该方法的检出限为10 3 至10 4 / ml。通过在cELISA之前溶解LPS可以达到这种敏感性。此外，这项研究中使用了一种新颖的基于免疫层析膜的手持分析（HHA）和PCR技术，针对图莱劳斯藻的17-kDa蛋白（TUL4）基因进行了靶向。与cELISA相比，HHA的灵敏度低约100倍，PCR的灵敏度高约10倍。所有这三种技术已成功应用于检测欧洲褐兔（Lepus europaeus）组织样品中的杜氏烟草。尽管所有感染的样品均通过cELISA识别，但细菌载量相对较低的样品却被PCR和HHA部分检测或未检测到，这可能是由于抑制剂或缺乏敏感性所致。总之，HHA可以用作执行现场诊断的非常快速和简单的方法，以获取土拉弗朗西斯氏菌感染的第一提示，尤其是在紧急情况下。在任何可疑情况下，均应通过cELISA和PCR等更敏感的技术来确诊。

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