解放军医学杂志2012，Vol.37Issue(5)：459-463,5.
Luminex液相芯片技术检测6种虫媒病毒方法的建立
Establishment of Luminex liquid chip technology for detection of six arboviruses
范丽 1李裕昌 2康晓平 2林方 2魏婧靖 2杨银辉 2熊正英1
作者信息
- 1. 710062西安 陕西师范大学体育学院
- 2. 100071北京 军事医学科学院微生物流行病研究所病原微生物生物安全国家重点实验室
- 折叠
摘要
Abstract
Objective To establish and evaluate liquid chip technique to detect six species of arbovirus including Tick borne encephalitis virus (TBEV), West Nile virus (WNV), Japanese encephalitis virus (JEV), Sindbis virus (SINV), Eastern equine encephalitis virus (EEEV) and Dengue fever virus (DENV). Methods Specific monoclonal antibodies for all of the six species of arbovirus were prepared and purified. The detecting antibodies were labeled with biotin and the capture antibodies were coupled to fluorescence coded microspheres. Then Luminex liquid chip technology was developed in a double antibody sandwich format. The singleplex and multiplex assay were conducted by measuring the virus cultures through Luminex 200 systems. Results A cutoff value twice higher than that of the background fluorescence was judged as positive reaction. The results of singleplex and multiplex assay for the virus cultures demonstrated specific positive signals, without cross reaction. Repeatability assays were also conducted and consentaneous results were gained, the coefficient of variability was less than 7% from inter and intra assay, implying that both the repeatability and stability of the array were good. The results demonstrated that using the developed Luminex liquid chip technology, the above arbovirus could be effectively identified and distinguished in single or in mixed. By applying series diluted virus cultures to this system, the detecting sensitivities for the six species of arbovirus were determined, which could reached 25pfu/ ml for TBEV, 781.2Spfu/ml for WNV, 781.25pfu/ml for JEV, 390.63pfu/ml for SINV, 781.2Spfu/ml for EEEV and 1562.5pfu/ml for DENV reached 781.2S, respectively. By comparing with the conventional ELISA, the liquid chip technique showed the advantages of higher sensitivity and repeatability, sample and time saving, and possessed the same specificity as ELISA. Conclusion A multiplex liquid chip for simultaneously detecting and serotyping six species of important arbovirus has been successfully developed, and could be potential useful for clinical virus detection.关键词
芯片分析技术/虫媒病毒/敏感性与特异性Key words
microchip analytical procedures/ arboviruses/ sensitivity and specificity分类
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范丽,李裕昌,康晓平,林方,魏婧靖,杨银辉,熊正英..Luminex液相芯片技术检测6种虫媒病毒方法的建立[J].解放军医学杂志,2012,37(5):459-463,5.
