两种块染技术对病毒感染细胞超薄切片染色效果的评价

Evaluation on the staining results of ultrathin sections of virus infected cells by two en bloc staining techniques

目的评价锇-硫代碳酰肼-锇(osmium-thiocarbohydrazide-osmium,OTO)及还原锇-硫代碳酰肼-锇(ferrocyanide-reduced-osmium-thiocarbohydrazide-osmium,ROTO)两种块染技术对病毒样品超薄切片的染色效果。方法收集四种病毒(人5型腺病毒、I型单纯疱疹病毒、人肠道病毒、呼吸道合胞病毒)感染的细胞样品,分别用OTO及ROTO技术进行块染,并以常规超薄切片染色为对照,通过透射电子显微镜观察并拍照,统计分析OTO及ROTO技术制备的病毒样品染色效果。结果OTO及ROTO法对病毒感染细胞样品的染色效果良好,病毒结构清楚,与常规染色效果相比无显著差异。结论OTO及ROTO技术可适用于多种类型病毒样品的块染处理,为病毒感染细胞样本的连续超薄切片、大尺度样品制备及电镜观测提供了依据。

Objective To evaluate the staining results of ultrathin sections of virus-infected cells by two en bloc staining techniques,i.e.osmium-thiocarbohydrazide–osmium(OTO)and ferrocyanide-reduced-osmium-thiocarbohydrazide-osmium(ROTO).Methods The samples of cells infected by four viruses(human adenovirus type 5,herpes simplex virus type 1,human enterovirus 71 and respiratory syncytial virus)were collected and stained by two en bloc staining methods,i.e.OTO and ROTO,respectively.Conventional staining method for ultrathin sections was used as the control.The samples were observed and photographed by transmission electron microscope(TEM)and the staining results of virus samples prepared by OTO and ROTO methods were analyzed statistically.Results The virus infected cell samples by OTO and ROTO methods showed good stained results and clear structures of the viruses,and the difference comparing to samples by conventional staining was not statistically significant(P>0.05).Conclusions Both OTO and ROTO methods can be applied as en bloc staining of virus infected cells samples and provided foundation for serial ultrothin sectioning,preparation of large-volume virus samples and for observation by EM.