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Injector based on a single cell of the microfluidic analytical system , solubilizing membranes, separation and active oxygen detection

Author: ZhangXinZuo
Tutor: ZhangLiSheng；TangBo；LiQingLing
School: Shandong Normal University
Course: Applied Chemistry
Keywords: Single Cell Analysis Microfluidic Chips Laser-induced fluorescence Lytic film Peroxide
CLC: Q257
Type: Master's thesis
Year: 2011
Downloads: 91
Quote: 0
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Abstract


Hydrogen peroxide (H 2 O 2 ) acts as a second messenger role in cellular signal transduction process, the degradation in the organism's physiology, pathology, organ, etc. plays a key regulatory role in the process. In addition, H 2 O 2 radicals and other reactive oxygen species such as superoxide anion (O 2 - . ) and hydroxyl radical (· OH) has a very close relationship. In the single-cell level H 2 O 2 detection for major diseases, early diagnosis, treatment, drug screening and cell physiology, pathological process has important guiding significance. This thesis is based on laboratory microfluidic chip to build their own single cells reactive oxygen analyzer and high-sensitivity laser-induced fluorescence detector based on the use of laboratory internal design and synthesis of a new type of green fluorescent probe bis (p-toluene sulfonyl the) - dichloro-fluorescein (FS), electric gantry injection method automation to achieve a single-cell injection, a single lytic film, efficient electrophoresis separation intracellular H the 2 O 2 Quantitative detection. This thesis consists of two parts, which reads as follows: The first chapter is the introduction, the main operating unit involved for many years single-cell component analysis, including single cell manipulation means derivative method, dissolved film means and detection means Overview, then a detailed overview of the microfluidic chip component analysis of single-cell, and explains the rise in recent years, high-throughput single-cell analysis technology. The second chapter to establish the the microfluidic injection method based on electric gantry system for the first time to achieve the quantitative detection of single HepG2 cells H 2 O 2 . The fluorescent probe FS incubated cells into the microfluidic chip sample cell, the use of electric gantry injection and laser-induced fluorescence detector automation in a simple cross chip to achieve a single-cell injection (cell sampling and single-cell capture) the H 2 O melting film, electrophoretic separation and single cell 2 quantitative detection. The method mainly includes three periods: the first period of (cell sample phase), the cells of the sample cell under the driving action of the electric field force flows through the cross intersection to the sample waste reservoir; second period (loading stage of the single-cell), changing the voltage condition will be the individual cells to be captured into the separation channel; third period (cell solubilizing membranes and electrophoretic separation stage), is captured in a single cell in the fast dissolving films, and electrophoretic separation occurs under the action of the high voltage electric field, and by the laser-induced fluorescence detected by the detector, while the other cells in the sample cell under the driving action of the electric field force is still flowing through the cross intersections, and flows into the sample waste pools, not interested in the separation and analysis of the cause interference. Single HepG2 cells detected by that method H 2 O 2 , the average content of 16.09 ± 9.84 amol (n = 15). The method is simple, fast, easy to implement, the advantages of automation and the degree of integration, the other components within a single cell provides a new approach for automated quantitative analysis.

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CLC: > Biological Sciences > Cell Biology > Cell physiology > Information transfer in cells
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