SBIR Phase II: Novel platform for the preparation of samples for cryo-TEM imaging: capturing biomacromolecules repeatably in vitreous ice without artifacts

小型企业创新研究(SBIR)第二阶段项目的更广泛影响是，通过解决相关工作流程中的主要瓶颈，为生命科学研究准备样本的新方法。这项拟议的工作提出了一种新的方法来制备生物样品，以便进行分析，以开发出清晰、高分辨率的分子3D模型。这将导致更快地获得更高质量的数据和更好的蛋白质和其他细胞成分的3D模型，为人类生物学提供新的见解，并改善人类健康。拟议的项目旨在解决目前在低温传输电子显微镜(CRYO-EM)样品制备方面遇到的最大问题。目前的样品制备技术是不可靠的，需要准备多余的网格来为生物大分子的3D模型获取足够的高质量数据。许多网格的质量很差，生成的数据不足以用于高分辨率蛋白质模型；或者样品可能包含空气/水界面相互作用产生的伪影，从而降低了最终分辨率。这一二期项目将推进一种创新的设计理念，利用无印迹样品制备技术重复地制备高质量样品。这项拟议的工作将探索贸易空间并优化图像分辨率的工艺参数，从而产生先进的原型。该奖项反映了NSF的法定使命，并已通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

The broader impact of this Small Business Innovation Research (SBIR) Phase II project is a new way to prepare samples for life sciences research by addressing the major bottleneck in the associated workflow. The proposed work advances a new method to prepare biological samples for analysis to develop crisp, high-resolution 3D models of molecules. This will result faster acquisition of higher quality data and better 3D models of proteins and other cellular components, providing new insight into human biology and improvements to human health. The proposed project aims to solve the largest issues currently experienced in cryogenic Transmission Electron Microscopy (cryo-EM) sample preparation. Current sample preparation technologies are unreliable and require preparation of excess grids to acquire enough quality data for a 3D model of a biomacromolecule. Many grids are of poor quality and generate insufficient data for a high-resolution protein model; or samples may contain artifacts from interaction at the air/water interface, reducing the ultimate resolution. This Phase II project will advance an innovative design concept to repeatably prepare high quality samples with a blotless sample preparation technique. The proposed work will explore the trade space and optimize process parameters for image resolution, leading to an advanced prototype.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.