Microfabricated fluorescence-activated cell sorters ([mu]FACS) for screening bacterial cells

cell sorter;flow cytometry;lab-on-chip;microfluidics

Fu, Anne Yen-Chen ; Quake, Stephen R. (co-advisor)

University:California Institute of Technology

Department:Chemistry and Chemical Engineering

关键词: cell sorter;    flow cytometry;    lab-on-chip;    microfluidics;   

Others  :  https://thesis.library.caltech.edu/1760/1/aycfuThesis.pdf

美国|英语

来源: Caltech THESIS

【 摘 要 】

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