STRUCTURE OF BUDDING VIRUS CORE PARTICLES

感染了Semlike森林病毒的BHK细胞正在研究中 使用半厚切片和IVEM。 这样做有两个目的 TRD项目。 第一种方法是在细胞中重建病毒核心颗粒， 组织培养细胞的细胞质。 二是要研究 核心颗粒的跨膜出芽过程。 我们认为这 一个TRD项目，因为我们正在使用病毒作为测试样本， 确定断层扫描可以获得的最大分辨率， 塑料型材 核心颗粒为40 nm，完整的 颗粒为70nm。 因此，可以肯定的是，大多数粒子不是 截短的，颗粒直径三倍数量级的部分 是必要的。 在以前的工作中，我们一直无法清楚地识别 细胞质中的核心颗粒，因为它们很少， 尽管可以看到细胞膜内和细胞膜上的出芽颗粒。 在过去的一年里，一种新的突变体被开发出来，其中病毒 细胞中的颗粒非常多，尽管没有看到出芽。 这样我们就有了所需的两种标本。 在以前的工作中， 我们无法获得足够的核心粒子的分辨率， 进行重建，大概是由于传统的标本， 制备和染色。 新材料由以下人员编写： 高压冷冻和冷冻置换，不使用 锇。 新材料的染色后切片看起来很有希望。 利用电子断层扫描技术研究了芽接过程， 直截了当。 我们现在集中精力确定我们是否有 足够的分辨率来进行旋转平均重建 核心粒子 切片的后染色可能会减少 分辨率或塑料嵌入本身可能会导致问题， 对比度匹配或粒度。 为了研究这个问题，我们有 布置成在200kV场发射TEM上记录一些测试图像， 能量过滤成像技术，在T博士的实验室里。S.纽约州立大学 奥尔巴尼物理系

BHK cells infected with the Semlike forest virus are being studied using semi-thick sections and the IVEM. There are two goals of this TRD project. The first is to reconstruct virus core particles in the cytoplasm of tissue culture cells. The second is to study the trans-membrane budding process of the core particle. We consider this a TRD project because we are using the virus as a test specimen to determine the maximum resolution that can be obtained by tomography on plastic sections. The core particles are 40nm and the complete particles are 70nm. Thus, to be sure most particles are not truncated, sections on the order of three times the particle diameter are needed. In previous work, we have been unable to clearly identify the core particles in the cytoplasm because they were very few, although budding particles in and on the cell membrane could be seen. Over the past year a new mutant has been developed, in which the virus particles are very numerous in the cells, although no budding is seen. Thus we now have the two types of specimen we need. In previous work, we could not obtain sufficient resolution of the core particles to make reconstructions, presumably due to conventional specimen preparation and staining. The new material has been prepared by high-pressure freezing and freeze-substitution, without the use of osmium. Post-stained sections of the new material looks promising. Study of the budding process by electron tomography is straightforward. We are now concentrating on determining if we have enough resolution to do rotationally-averaged reconstructions of the core particles. Post-staining of the sections may decrease the resolution, or plastic embeddment itself might cause a problem by contrast-matching or granularity. To investigate this issues, we have arranged to record some test images on a 200kV field-emission TEM with energy-filtered imaging, which is in the lab of Dr. T. S. Kuan of SUNY Albany Physics Dept.