Biotechnology and Bioprocess Engineering Applications of Raman Spectroscopy

改进的过程分析技术(PAT)的发展是学术和工业工程师的一个重要的研发目标，也是一个日益需要跨学科方法的目标。我实验室的研究主要集中在开发基于拉曼光谱(RS)的生物技术/生物过程应用的分析方法和技术。不同形式的RS是光学非弹性散射技术，它直接探测分子的振动状态，从而提供与红外吸收光谱相似的信息，但在确定水环境中生物分子的浓度和构象方面具有独特的优势。RS可以使用不干扰活细胞的波长和强度来实现，并且可以与显微镜平台相结合，使用分子振动作为对比机制来提供细胞的无标记化学图像。这些空间分辨的化学图像可以提供关于细胞周期、表型、活性和新陈代谢的丰富信息。我的实验室在遥感仪器方面的最新技术进步进一步允许使用一种称为受激拉曼光谱(SRS)的技术来近实时地解析细胞特征的变化。这项建议描述了未来三个相关方向的研究活动，利用RS和SRS：(1)开发一种非破坏性方法，用于在干细胞衍生的基于细胞的疗法的制造过程中表征和监测细胞，该方法提供新的QC过程分析技术，利用也提供正交分析数据的原位光学方法来增强传统的(破坏性的)采样细胞分析；(2)开发允许血库和医院评估与存储相关的降解，从而评估密封储存袋中的高价值血液产品是否适合输血的仪器和方法；(3)开发分析工具，使生化工程师能够更有效地筛选细胞培养物，以在生物过程开发期间为高产细胞生产特定蛋白质，并更好地了解和监测生产过程中降低生产率的细胞变化。这些是我实验室中新项目的例子，这些项目构成了一个持续研究计划的核心，该计划旨在开发基于RS和SRS的有价值的新光谱工具，用于生物技术和生物过程工程中的应用。

The development of improved process analytical technologies (PAT) is a crucial R&D objective for both academic and industrial based engineers and one that increasingly demands an interdisciplinary approach. Research in my laboratory is focused mainly on development of analytical methods and technologies for biotechnology/bioprocess applications based on Raman spectroscopy (RS). The various forms of RS are optical inelastic scattering techniques that directly probe the vibrational states of molecules and thus provide similar information as infrared absorption spectroscopy, but offer distinct advantages for determining the concentrations and conformations of biomolecules in aqueous environments. RS can be implemented using wavelengths and intensities that are non-perturbing to live cells, and can be coupled with a microscopy platform to provide label-free chemical images of cells using molecular vibrations as a contrast mechanism. These spatially resolved chemical images can provide a wealth of information about cell cycles, phenotypes, viability and metabolism. Recent technological advances in RS instrumentation available in my laboratory further allows near real-time temporal resolution of changes in cellular characteristics using a technique called stimulated Raman spectroscopy (SRS). This proposal describes future research activities in three related directions that exploit both RS and SRS: (1) to develop a non-destructive methodology for characterizing and monitoring cells during the manufacturing of cell-based therapeutics derived from stem cells which provides new QC process analytical technologies that augment conventional (destructive) sampled cell analyses with an in situ optical approach that also offers orthogonal analytical data; (2) to develop instrumentation and methods that allow blood banks and hospitals to assess storage-related degradation, and thus the suitability for transfusion, of high-value blood products within sealed storage bags; (3) to develop analytical tools that allow biochemical engineers to more efficiently screen cultures of cells engineered to produce specific proteins for high-producing cells during bioprocess development, and to better understand and monitor cellular changes during production that degrade productivity. These are examples of new projects in my laboratory that comprise the core of a continuing research program directed toward developing valuable new spectroscopic tools based on RS and SRS for applications in biotechnology and bioprocess engineering.