Ultra-high Throughput, High Purity Stem Cell Isolation

描述（由申请人提供）：该项目的目标是开发一种非常高速（10亿个细胞/小时）的细胞分选系统，该系统具有分层结构，使用多种标记来获得来自成人外周血或骨髓的超高纯度（99.999%）造血干细胞（HSC）样本及其来自成人骨髓的前体间充质干细胞（MSC）样本。来源于成人的造血干细胞/间充质干细胞避开了围绕胚胎干细胞使用的伦理问题，同时为糖尿病、帕金森病、脊髓损伤等疾病的细胞替代疗法带来了希望。本项目第一期的具体目标是：1。开发高速细胞分选器的总体架构，包括光学、数据采集和控制子系统。2. 论证了单通道切换速度达到2000个单元/秒的可行性。3. 开发微加工并行通道阵列芯片Gigasort的设计，实现10亿个单元/小时的分选速度。4. 制定Teragenics高速细胞分选仪的测试和验证计划，包括用于特定细胞标记物的初步实验方案。所提出的细胞分选器的核心包括一个微流控芯片Gigasort （Teragenics正在申请专利），它利用快速切换方法以及高度并行性来实现高速细胞分选，并采用分层方法来实现高分选纯度。Gigasort芯片采用大批量微加工技术制造，成本低，而且是一次性的。Gigasort芯片与用于光学检测和读出，控制电子，试剂存储等的仪器子系统接口，并可用于HSC/MSC的多种阳性和阴性标记，如CD34， CD34+, Sca+， lin -在单一层次排序中。拟议的项目将提供高速细胞分选器的蓝图，该分选器比现有方法更快地提供3个数量级的HSC/MSC纯度，其成本模型允许Gigasort微流控芯片一次性使用，从而为临床应用保持完全无菌的环境。

DESCRIPTION (provided by applicant): The goal of this project is to develop a very high-speed (1 billion cells/hour) cell sorting system with hierarchical architecture using multiple markers to achieve ultra high purity (99.999 percent) Hematopetic Stem Cells (HSC) samples derived from adult peripheral blood or bone marrow and their precursor Mesenchymal Stem Cell (MSC) samples derived from adult bone marrow. HSC/MSC derived from adult sources circumvent the ethical issues surrounding the use of embryonic stem cells while holding the promise for cell replacement therapies for diseases such as diabetes, Parkinson's Disease, spinal chord injury, and many others. Specific aims of Phase 1 of this project are: 1. Develop overall architecture of the high-speed cell sorter including the sub-systems for optics, data acquisition and control. 2. Demonstrate the feasibility of single channel switching speed of 2,000 cells/second. 3. Develop the design of micro-fabricated parallel channel array-chip, Gigasort, to achieve sorting speed of 1 billion cells/hour. 4. Develop a plan for test and validation of Teragenics' high-speed cell sorter including preliminary experimental protocols for use with specific cell markers. The core of the proposed cell sorter comprises of a microfluidic chip, Gigasort (Teragenics patent pending), which takes advantage of a rapid switching methodology coupled with a high degree of parallelism to enable the cell sorting at high speeds and a hierarchical approach to sorting to enable high sorting purity. The Gigasort chips are manufacturable using high volume microfabrication techniques and are low cost and disposable. The Gigasort chip interfaces with instrument sub-systems for optical detection and readout, control electronics, reagent storage etc and works with multiple positive and negative markers for HSC/MSC such as CD34, CD34+, Sca+, Lin-in a single hierarchical sort. The proposed project will deliver the blue print for the high speed cell sorter that delivers 3 orders of magnitude purer HSC/MSC samples 4 times faster than available methods with a cost model that allows the Gigasort microfluidic chip to be disposable, thus maintaining a fully sterile environment for clinical applications.