COLUMN FRACTION SCREENING OF CPMV PARTICLES

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 需要能够分离和表征UV处理后的RNA填充、空和聚集部分，以便提纯包含RNA的、非感染性的单个CPMV颗粒，用于进一步的体内研究。为了显示许多柱级分是否具有这些物理特性，目标是能够以有效的方式可视化从柱子或蔗糖梯度中收集的级分。由于一次典型的精馏分产率在20-100个馏分之间，因此为EM制备单独的CPMV馏分将从自动化中受益匪浅。然后需要分析透射电子显微镜图像，以查看每个组分中的颗粒，以识别和量化RNA填充的衣壳、空衣壳、聚集碎片和破碎颗粒的比例。这一程序将使个体灭活的大量CPMV纳米颗粒的验证在体内使用是可行的。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. There is a need to be able to separate and characterize the RNA-filled, empty, and aggregated fractions following UV treatment in order to purify the RNA-containing, non-infectious single CPMV particles for further in vivo studies. In order to show whether the many column fractions have these physical characteristics, the goal is to be able to visualize fractions collected from columns or sucrose gradients in an efficient manner. Since a typical column run yields between 20-100 fractions, the preparation of individual CPMV fractions for EM would benefit significantly from automation. Analysis of the TEM images would then be needed, to view the particles in each fraction to identify and quantify the proportion of RNA-filled capsids, empty capsids, aggregated debris, and broken particles. This procedure would make the validation of individual inactivated lots of CPMV nanoparticles feasible for in vivo use.