Wang Wei, Butler Eboneé N, Veguilla Vic, Vassell Russell, Thomas J Terrig, Moos Malcolm, Ye Zhiping, Hancock Kathy, Weiss Carol D
Division of Viral Products, Center for Biologics Evaluation and Research, US Food and Drug Administration, 1401 Rockville Pike, Rockville, MD 20852, USA.
J Virol Methods. 2008 Nov;153(2):111-9. doi: 10.1016/j.jviromet.2008.07.015. Epub 2008 Sep 4.
Pseudotype reporter viruses provide a safe, quantitative, and high-throughput tool for assessing antibody neutralization for many viruses, including high pathogenicity H5 and H7 influenza A strains. However, adapting this system to other influenza subtypes has been hampered by variations in the protease cleavage site of hemagglutinin (HA) that make it less susceptible to the cleavage required for infectivity. In this report several proteases, reporter vectors, and cell substrates were evaluated while optimizing pseudovirus production, and robust methods were established for sensitive and specific neutralization of pseudotypes carrying influenza H1, H3, and H5 subtype HA that correlates well with titers obtained in microneutralization assays involving replicating influenza virus These findings should facilitate broad use of HA-pseudotypes that remove the need for infectious virus in a range of applications, including neutralization assays for serological surveys of viral infection and evaluations of vaccine sera.
假型报告病毒为评估包括高致病性H5和H7甲型流感病毒株在内的多种病毒的抗体中和作用提供了一种安全、定量且高通量的工具。然而，由于血凝素（HA）蛋白酶切割位点的变异使得其对感染所需的切割不太敏感，将该系统应用于其他流感亚型受到了阻碍。在本报告中，在优化假病毒生产的同时评估了几种蛋白酶、报告载体和细胞底物，并建立了稳健的方法用于对携带流感H1、H3和H5亚型HA的假型进行敏感且特异的中和，这与在涉及复制流感病毒的微量中和试验中获得的滴度高度相关。这些发现应有助于广泛使用HA假型，从而在一系列应用中无需使用感染性病毒，包括用于病毒感染血清学调查的中和试验以及疫苗血清的评估。
