Development of the rapid diagnosis test for avian influenza viruses

由禽流感病毒突变引起的大流行性流感的爆发一直是世界上令人担忧的问题。当大流行性流感爆发时，必须采用免疫层析快速诊断试验，将大流行性流感与季节性流感和其他发热性疾病区分开来。为了建立快速诊断检测方法，制备了一种单克隆抗体4E3，该抗体与a /Turkey/Ontario/7732/66(H5N9)（免疫原）NP n端1/3位表位发生反应，并用于免疫层析（夹心法）固相抗体和彩色乳胶偶联抗体。该方法在10 min内检出A/Crow/Kyoto/04（H5N1）病毒至少4 × 10^5的病灶形成单位(FFU)/ml，在H5N2、H5N3、H5N9中分别检出3.4 × 10^5、5.3 × 10^5、1.1 × 10^5FFU/ml。与除H5病毒外的15株（H3 ~ H15）禽流感病毒（AIV）均有反应。另一方面，它与19种人类流感病毒（A型和B型）、22种其他呼吸道病毒和24种细菌都没有反应。此外，2004年在泰国从10名患者身上分离出的10株AIV（H5N1， Z型，进化枝1）均被快速检测出，2006年在印度尼西亚从1名患者身上分离出的1株AIV（H5N1，可能进化枝2-1）中有1株被快速检测出，从2头病猪身上分离出的2株H5N1病毒中有2株被快速检测出，从7只死鸡身上分离出的7株H5N1病毒中有7株被快速检测出。目前，禽流感病毒感染患者的鼻咽区H5N1病毒载量似乎不像季节性流感病例那样高。然而，一旦在人类之间具有高度传染性的大流行性流感病毒出现，病毒就可能在鼻咽区充分传播，达到季节性流感的水平。结果表明，该快速诊断方法具有足够的敏感性、特异性、抗原识别的稳定性和快速性能。这种检测应该显示出强大的能力，作为快速遏制和分类的有力工具，特别是在大流行性流感的早期阶段。少

The outbreak of a pandemic influenza derived from a mutated avian influenza virus has been an apprehensive issue in the world. When the pandemic influenza broke out, a rapid diagnosis test by immunochromatography should be indispensable to distinguish the pandemic from the seasonal influenza and other febrile illness. To establish the rapid diagnosis test a monoclonal antibody, 4E3, which reacts with an epitope located in 1/3 of N-terminus of NP of A/Turkey/Ontario/7732/66(H5N9) (immunogen) was produced and used for both solid-phase and colored latex-conjugated antibody of an immunochromatography(sandwich method). This rapid test detected at least 4x10^5 focus forming unit(FFU)/ml of A/Crow/Kyoto/04(H5N1) in 10 min, and it detected 3.4x10^5, 5.3x10^5, 1.1x10^5FFU/ml of an H5N2, H5N3, H5N9 virus, respectively. It also reacted with 15 strains(H3〜H15) of avian influenza viruses(AIV) except H5 viruses. On the other hand it neither reacted with 19 human influenza viruses(A and B type), 22 o … More ther respiratory viruses or 24 bacteria. Furthermore, the rapid test detected all 10 strains of AIV(H5N1, Z type, clade 1) isolated from 10 patients in 2004 in Thailand and it also detected 1 of 1 strain of AIV(H5N1, probably clade 2-1) isolated from one patient, 2 of 2 strains of H5N1viruses isolated from 2 ill pigs, and 7 of 7 strains of H5N1 viruses isolated from 7 dead chickens in 2006 in Indonesia. At present H5N1 viral load at nasopharyngeal region in patients with AIV infections seems not high as in case of seasonal influenza. However, once the pandemic influenza viruses emerged which is very contagious among humans, the viruses could propagate sufficiently in the nasopharyngeal region to the level of seasonal influenza. These results reveal that this rapid diagnosis test has enough sensitivity, specificity, stability for antigen recognition and rapid performance. This test should display great ability as a powerful tool for the rapid containment and triage especially in the early-phase of the pandemic influenza. Less