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首页> 外文学位 >Microfabricated fluorescence-activated cell sorters (muFACS) for screening bacterial cells.
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Microfabricated fluorescence-activated cell sorters (muFACS) for screening bacterial cells.

机译：用于筛选细菌细胞的微型荧光激活细胞分选仪（muFACS）。

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In this thesis, I have developed elastomeric microfabricated cell sorting devices using a micromachining technology, “soft lithography”. Inexpensive elastomeric microfabricated devices were designed to replace flow chambers in conventional fluorescence-activatived cell sorters (FACS). Sorting of cells and other particles was accomplished via different means of flow control. My early work of cell sorting on these devices was accomplished using electrokinetic flow. However, in order to alleviate problems associated with electrokinetic flow, the microfabricated cell sorter was integrated with microvalves and micropumps for pneumatic actuation control. The integrated cell sorter has better capabilities of fine-tuning the flow control, manipulating single cells and is less harmful to the cells than electrokinetic flow. Substantial enrichments of beads and cells were accomplished on these devices. Novel sorting algorithms, which can only be implemented in microfabricated devices, were also demonstrated. Compared with conventional FACS, these microfabricated devices allow for more sensitive optical detection for bacterial cells and DNA, innovative sorting schemes and are disposable to eliminate any cross-contamination from previous runs. Ultimately, these elastomeric microfluidic flow cells provide an inexpensive, robust and effective way to perform cell sorting and can be used as stand-alone devices or as a part of an integrated system for diagnostics and/or cytometric measurements.; Presently, the microfabricated cell sorter is enjoying new applications in various fields for high throughput screening, including directed evolution, digital genetic circuits, microbiology and cell biology of gene expression and regulation. In addition, this sorter is not limited only to the detection of optical signals. I have attached the sorter to a high resolution magnetometer, a superconducting quantum interference device (SQUID) microscope, to obtain cytometric data of the magnetic field strengths of magnetotactic bacteria as they flowed through the device.; This thesis lays down the foundation for future work in cell sorting and single cell analysis. Time-course measurements of a single cell for kinetic studies can be implemented using novel sorting schemes. Sample dispensing and any downstream analysis, such as cell lysis and/or polymerase chain reaction, can be carried out immediately after the cells have been sorted. The sorter could also be incorporated with other technologies to measure cellular magnetic or electrical properties. We anticipate that an integrated lab-on-chip, where cell sorting is one of the steps of a complete analysis system, is not far off.
机译：在本文中，我开发了一种使用微加工技术“软光刻”的弹性体微细细胞分选设备。廉价的弹性体微细加工设备设计用于替代常规荧光激活细胞分选仪（FACS）中的流动室。细胞和其他颗粒的分选是通过不同的流量控制手段完成的。我在这些设备上进行细胞分选的早期工作是使用电动流完成的。然而，为了减轻与电动流动相关的问题，将微型细胞分选机与微型阀和微型泵集成在一起以进行气动控制。集成式细胞分选仪具有更好的微调流量控制，操纵单个细胞的功能，并且与电动流动相比，对细胞的危害较小。在这些装置上完成了珠子和细胞的大量富集。还展示了只能在微型设备中实现的新型排序算法。与传统的FACS相比，这些微型设备可对细菌细胞和DNA进行更灵敏的光学检测，创新的分选方案，并且可一次性使用，以消除先前运行中的任何交叉污染。最终，这些弹性体微流体流通池提供了一种廉价，坚固而有效的方式来进行细胞分选，并且可以用作独立设备或用作诊断和/或细胞计数测量的集成系统的一部分。目前，微细加工细胞分选仪在高通量筛选的各个领域中都有新的应用，包括定向进化，数字遗传电路，微生物学以及基因表达和调控的细胞生物学。另外，该分类器不仅限于光信号的检测。我已经将分选机连接到高分辨率磁力计，超导量子干扰设备（SQUID）显微镜上，以获取趋磁细菌流经该设备的磁场强度的细胞数据。本文为细胞分选和单细胞分析的未来工作奠定了基础。可以使用新颖的分类方案对单个细胞进行动力学研究，进行时程测量。样品分选后，可以立即进行样品分配和任何下游分析，例如细胞裂解和/或聚合酶链反应。该分选机还可以与其他技术结合使用，以测量细胞的磁性或电学性质。我们预计，将细胞分选作为完整分析系统的步骤之一的集成芯片实验室也不远。

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