An intergrated analytical microfluidic culture system for stem cell research

干细胞提供了在培养中生成几乎所有身体组织类型的可能性，最近干细胞生物学的重大进展引发了新的研究途径，最终将使临床医生能够利用这种治疗潜力来革新再生医学。然而，目前很难可靠地产生足够数量的这些细胞，主要是因为这些总是混合培养的群体在体外倾向于以非定向的方式分化。这大大减缓了实现这些细胞的全部临床前景的进展。最近，已经有可能设计出微型组织培养芯片，允许数千个单细胞在独立控制的介质条件下生长并同时进行分析，实际上允许数百个平行实验，而成本只是单个大规模实验的一小部分。实现这项技术非凡潜力的一个主要挑战是，由于培养体积小，每个实验只有很少的细胞，监测细胞的局部环境以及它们的代谢和分化状态极其有限。我们建议通过设计一种基于荧光和拉曼光谱成像的完全无损、非接触的光学分析系统来解决这些挑战，该系统允许对细胞及其环境进行快速、实时、无损的原位分析。我们组建了一支在每一项关键技术-干细胞培养、微流控和光谱学-方面拥有专业知识的科学家和工程师团队，以实现和开发这一集成分析微流控培养系统的独特能力。这里提出的研究的主要目标是详细分析和优化控制单个干细胞分化和扩增的因素，这些因素在目前的培养系统中是不可能的。这项研究将产生关于关键培养条件和其他限制这些细胞在培养中扩张和受控分化的因素的宝贵信息。这项工作的结果将导致大规模生产干细胞的创新新方法，并将加快和降低开发重要的基于干细胞的新疗法的成本。

Stem cells offer the potential to allow virtually every tissue type in the body to be generated in culture and major recent advances in stem cell biology have sparked new avenues of research that will ultimately allow clinicians to harness this therapeutic potential to revolutionize regenerative medicine. Presently, however, it is difficult to reliably produce these cells in sufficient numbers, principally because these invariably mixed culture populations tend to differentiate in a non-directed manner in vitro. This greatly slows progress toward realizing the full clinical promise of these cells. Very recently, it has become possible to engineer micro-scale tissue culture "chips" that allow thousands of single cells to be grown under independently controlled media conditions and analyzed simultaneously, in effect allowing hundreds of parallel experiments at a fraction of the cost of a single large-scale experiment. A major challenge in realizing the extraordinary potential of this technology is that monitoring the local environments of the cells, as well as their metabolic and differentiation state, is extremely limited due to the small culture volumes and few cells per experiment. We propose to solve these challenges by engineering a completely nondestructive, non-contact, optical analytical system, based on fluorescence and Raman spectroscopic imaging, that permits fast, real-time, nondestructive, in situ analysis of both the cells and their environment. We have assembled a team of scientists and engineers with expertise in each of the key technologies-stem cell culture, microfluidics and spectroscopy-to realize and exploit the unique capabilities of this integrated analytical microfluidic culture system. The primary goal of the research proposed here is to analyze, in fine detail, and optimize the factors that control the differentiation and expansion of single stem cells in ways not possible with current culture systems. This research will yield invaluable information about critical media conditions and other factors that limit the expansion and controlled differentiation of these cells in culture. The results from this work will lead to innovative new approaches for large-scale production of stem cells and will accelerate and reduce the cost of developing important new stem-cell-based therapies.