Suzuki T, Jyono M, Tsukimoto M, Hamaoka A, Suzuki Y
Department of Biochemistry, University of Shizuoka, School of Pharmaceutical Science, Japan.
Anal Biochem. 1995 Jun 10;228(1):42-7. doi: 10.1006/abio.1995.1312.
A procedure for labeling influenza viruses with alkylamine-modified horseradish peroxidase (HRP) was developed to study virus receptor specificities in a virus overlay assay. Six alkylamine derivatives of HRP were prepared by the periodate oxidation method. Human influenza virus A/PR/8/34 (H1N1) was labeled with the alkylamine derivatives, and a variety of parameters for labeling were tested in a fetuin-immobilized microplate solid-phase assay. The labeling efficiency of influenza virus with the derivatives of HRP was affected by the alkyl chain length of the derivatives. The most effective derivative for labeling the virus being tested was octylamine-modified HRP (octyl-HRP). The labeling profile of A/PR/8/34 by octyl(C8H17)-HRP was analyzed b sucrose density gradient centrifugation. The labeling had no effect on viral hemagglutinating activity and fetuin binding specificity. The specific binding of the HRP-labeled human influenza viruses (A/PR/8/34 and B/Lee/40) to glycolipids was demonstrated by a virus overlay assay. In this virus-HRP-labeled system, the virions which bound to gangliosides developed on a silica gel thin-layer plate were detected simply and rapidly.
开发了一种用烷基胺修饰的辣根过氧化物酶（HRP）标记流感病毒的方法，以在病毒覆盖试验中研究病毒受体特异性。通过高碘酸盐氧化法制备了六种HRP的烷基胺衍生物。用人流感病毒A/PR/8/34（H1N1）对这些烷基胺衍生物进行标记，并在胎球蛋白固定的微孔板固相试验中测试了各种标记参数。HRP衍生物对流感病毒的标记效率受衍生物烷基链长度的影响。用于标记被测病毒的最有效衍生物是辛胺修饰的HRP（辛基-HRP）。通过蔗糖密度梯度离心分析了辛基（C8H17）-HRP对A/PR/8/34的标记情况。该标记对病毒血凝活性和胎球蛋白结合特异性没有影响。通过病毒覆盖试验证明了HRP标记的人流感病毒（A/PR/8/34和B/Lee/40）与糖脂的特异性结合。在这个病毒-HRP标记系统中，在硅胶薄层板上发育的与神经节苷脂结合的病毒粒子能够被简单快速地检测到。
