快速分子检测和流感病毒A和B的分化
Summary
我们描述了一个快速，分子为基础的甲型和乙型流感检测。流感检测采用等温扩增与流感特异性引物，其次是目标检测与分子信标探针检测15分钟内每一个目标。的甲型和乙型流感检测是用户友好的，并要求最少的动手时间来执行。
Abstract
流感是在人中由流感病毒引起A和B的传染性呼吸系统疾病，并导致每年发病率和死亡率的一个显著量。的甲型和乙型流感检测是第一个可用的CLIA-放弃分子快速流感测试。流感A和B的测试工作通过采用等温扩增与流感特异性引物，接着目标检测的分子信标探针。这里，A型流感的性能和B测定冷冻，归档鼻咽拭子（NPS）试样储存在病毒运输介质（VTM）进行比较，以呼吸面板测定。
流感A和B测定的性能通过的结果相比较，以呼吸面板参考方法进行评价。总甲型流感病毒的敏感性为67.5％（95％CI（CI），56.6-78.5），特异性为86.9％（CI，71.0-100）。对于乙型流感病毒的检测，其敏感性和特异性分别为90.2％（CI，68.5-100）和98.8％（CI，68.5-100）表示。
该系统具有的显著缩短测试时间比其他任何现有的分子检测及简单，无吸管程序运行的优势一个完全集成的，封闭的，小尺寸的系统。总体而言，流行性感冒A和B测定本研究评估具有作为一个点 – – 关心快速流感诊断测试的可能性。
Introduction
流感病毒感染导致发病率和死亡率，每年1，2，3的显著量。单纯性流感的特点是宪法和呼吸道症状，如发热，肌痛，头痛和非生产性咳嗽4,5。老年人，幼儿，免疫功能低下患者，和患者的潜在合并症处于严重的并发症，如肺炎，心肌炎，中枢神经系统疾病，甚至导致死亡6,7风险较高。
流感感染是抗病毒治疗的症状出现后48小时内及时服用能减少疾病的严重程度和长度从8其他呼吸道病毒唯一的。流感的快速鉴定也已示出减少使用的不必要的抗生素9,10。另外，住院患者感染流感病毒必须放置在适当的感染控制预防措施隔离室。然而，所造成的非流感病毒呼吸道疾病可能难以从流感临床区分。出于这个原因，对于流感快速准确的诊断测试是用于临床患者管理非常重要。
数测定法可用于检测和流感病毒鉴定。快速流感抗原检测试验（RIDTs）被广泛应用于临床实践中作为点护理测试，因为它们易于使用，并提供在15至30分钟11，12的结果;然而，他们的敏感性有很大的不同取决于制造商和人口（10-80％）进行测试，并且流感类型和subtyPE 13，14，15。直接荧光测定（的DFA）提供超过RIDTs优异灵敏度，但处理时间大于（〜3小时），并且必须由熟练的技术人员16，17来完成。病毒培养一直是流感的诊断的金标准，并已改善了两个RIDTs和DFA的18灵敏度。然而，流感病毒培养可以从2-14ð任何地方来完成，减少在帮助病人管理19其效用。最后，核酸扩增试验（NAAT）已经取代培养技术在流感诊断新的金标准。 NAAT被认为具有在几个小时检测流感的最大灵敏度。然而，NAAT是最昂贵的测定和需要专门的设备和技术来执行5 <s了>，20，21，22，23，24，25。
此处所描述的流感A和B的测定法是第一CLIA-放弃分子快速流感测试是容易获得的。此法的工作原理是采用使用特定流感引物，其次是目标检测与分子信标探针等温扩增核酸内切酶泥炭截短扩增反应（NEAR）。该测定区分流感A选自B，需要2分钟来设置和处理一个样品，并且总共需要15分钟来完成。
这里，我们目前的流感A＆B试验的协议。此外，我们还提供了一个样本数据集比较流感A对已归档的鼻咽拭子（NPS）的性能和B检测标本保存在病毒的传输介质（VTM）到其他呼吸道病原体面板检测。
Protocol
Representative Results
Discussion
流感病毒是发病率和死亡率的显著世界范围的原因。流感的快速和准确的诊断是主要键期间呼吸季节管理流感爆发中的一个。其他基于抗原的测试是快速和容易执行;然而，它们具有低的灵敏度13。另一方面，传统的分子测试具有改善的敏感性，但需要更多的有经验的化验师来执行，而且更昂贵。在这项研究中和协议中描述的流感A和B测定是流感A和B的CLIA-放弃，快速的分子测试的?…
Declarações
The authors have nothing to disclose.
Acknowledgements
We thank the Clinical Microbiology Service staff of the Memorial Sloan-Kettering Cancer Center for help in collecting clinical specimens. This study was supported in part by a research agreement between MSKCC and Alere Scarborough (SK2013-0262).
Materials
| Alere i Instrument | Alere | NAT-000 (Global), NAT-024 (US) | |
| Alere i Influenza A & B 24 Test Kit | Alere | 425-000 (Global), 425-024 (US) | |
| Alere i Barcode Scanner | Alere | EQ001001 | |
| Alere Universal Printer | Alere | 55115 (Global), alereiprinter (US) | |
| 200 µL precision pipette | |||
| 200 µL disposable pipette tips | |||
| Viral transport medium | Remel | M4-RT |
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