Abstract：
The generation and release of mature virions from human cytomegalovirus (HCMV) infected cells is a multistep process, involving a profound reorganization of cellular structures and various stages of virus particle morphogenesis in different cellular compartments. Although the general steps of HCMV morphogenesis are known, it has become clear that the detailed molecular mechanisms are complex and dependent on various viral factors and cellular pathways. The lack of a full understanding of HCMV virion morphogenesis emphasizes the need of imaging techniques to visualize the different stages of virion assembly, such as electron microscopy. Here, we describe various electron microscopy techniques and the methodology of high-pressure freezing and freeze substitution for sample preparation to visualize HCMV morphogenesis. These methods are used in our laboratory in combination with a thorough quantification to characterize phenotypic alterations and to identify the function of viral and cellular proteins for the various morphogenesis stages.
摘要：
从人巨细胞病毒（HCMV）感染的细胞中产生和释放成熟的病毒体是一个多步骤的过程，涉及细胞结构的深刻重组和不同细胞区室中病毒颗粒形态发生的不同阶段。虽然HCMV形态发生的一般步骤是已知的，很明显，详细的分子机制是复杂的，并且依赖于各种病毒因子和细胞途径。缺乏对HCMV病毒体形态发生的充分了解强调需要成像技术来可视化病毒体组装的不同阶段，比如电子显微镜。这里,我们描述了各种电子显微镜技术以及高压冷冻和冷冻替代样品制备的方法，以可视化HCMV形态发生。这些方法在我们的实验室中与彻底的定量结合使用，以表征表型改变并鉴定病毒和细胞蛋白质在各种形态发生阶段的功能。
