| Single-cell analysis is of significant interest to the biological, medical, and pharmaceutical communities, because it is essential to a better understanding of basic cellular functions and it can also provide information about the cell-to-cell variation in large populations of cells. Recently the exploitation of microfluidic chip-based systems for biological cell studies is attracting broad interest. Such systems, more generally referred to as miniaturized total analysis systems (μTAS), had gone through rapid development in the past decade. The micrometer channel dimensions of microfluidic chips are ideally suited for the sample introduction, manipulation, reaction, separation and detection of single cells. It has significant academic and practical value to develop simple, fast, sensitive methods on the basis of these analysis techniques. The research work was carried out to focus on the mentioned points. The dissertation, a collection of the research results, contains the following six chapters.In the chapter one, first of all, the principle of microfluidic chip was introduced briefly. Then the application of microfluidic chip in the single-cell analysis was reviewed in detail. Finally, the work for microfluidic chip with electrochemiluminescence (ECL) detection was summarized.In the chapter two, a new method for the detection of taurine was developed using capillary electrophoresis by derivatization of 2,3-naphthalene-dicarboxaldehyde (NDA) pre-column. The effects of the derivatization time, detected potential, running buffer pH, running buffer concentration and separation voltage were studied. The optimum conditions of separation and detection were 8.00×10-3 mol·L-1 Na2B4O7 (pH 11.04) for the buffer solution, 20 kV for the separation voltage, and 0.9 V (vs. SCE) for the detection potential. The limit of detection is 1.60×10-6 mol·L-1 (S/N = 3) and the linear range is 5.00×10-6-1.00×10-3 mol·L-1 with a correlation coefficient of 0.9997 for the injection voltage of 5 kV and the injection time of 10 s. The response for a series of seven injections of 1.00×10-3 mol·L-1 taurine resulted in a relative standard deviation of 1.7% for the migration time, and 2.5% for the electrophoretic peak current, respectively. The method was used to detected taurine both in normal hemolysis and abnormal hemolysis. The recovery of the method was between 93-95%.In the chapter three, a PDMS/glass microfluidic chip with integrated electrochemical detection cell equipped by replaceable micro-disk working electrode was developed. The end-channel amperometric detector contained a guide tube for alignment of working electrode, which made the alignment between carbon fiber microdisk working electrode and separation channel done precisely without the help of a micro-positioner. Using ascorbic acid (AA) and dopamine (DA) as the model analytes, the performance of the fabricated PDMS/glass microfluidic chip was characterized. The experiment results indicate that the alignment of the electrode to the channel outlet can be carried out accurately and quickly. Good reproducibility of the electrode alignment was demonstrated by a 3.1% RSD for the electrophoretic peak current of AA obtained after repeatedly position of a micro-disk working electrode.In the chapter four, the self-assemble microfluidic chip electrochemical detection system (microfluidic chip-ED) was employed for the analysis of ascorbic acid in rat peritoneal mast cells lysis with a carbon fiber microdisk bundle electrode (CFMBE). The optimum conditions of separation and detection were 1.0×10-2 mol·L-1 borate buffer (pH 9.0) for the buffer solution, 1.1 kV for the separation voltage, and 0.8 V (vs. Ag/AgCl) for the detection potential, The limit of detection is 2.7×10-6 mol·L-1 (S/N = 3) and the linear range is 8.0×10-6-5.0×10-4 mol·L-1 with a correlation coefficient of 0.9985 for the injection voltage of 1.0 kV and the injection time of 5 s. The response for a series of seven injections of 5.0×10-4 mol·L-1 AA resulted in a relative standard deviation of 0.57% for the migration time, and 2.1 % for the electrophoretic peak current, respectively. The method was used to detect ascorbic acid in rat peritoneal mast cells lysis. The recovery of the method was between 94-97%.In the chapter five, a kind of novel CFMBE with double modifiers - multiwalled carbon nanotubes (MWCNTs) and poly(diallyldimethylammonium chloride) protected Prussian blue nanoparticles (P-PB) in microfluidic chip-ED was applied to determine AA in individual rat peritoneal mast cells. The catalytic electrochemical properties of MWCNTs/P-PB CFMBE could enhance sensitivity obviously compared with CFMBE at a relatively lower detection potential (+0.4 vs. Ag/AgCl). Under the optimum conditions of detection, the limit of detection is 2.7×10-7 mol·L-1 (S/N = 3) and the linear range is 8.0×10-7-5.0×10-4 mol·L-1 with a correlation coefficient of 0.9981. The response for a series of seven injections of 5.0×10-5 mol·L-1 AA resulted in a relative standard deviation of 0.86% for the migration time, and 2.3% for the electrophoretic peak current, respectively. A single rat peritoneal mast cell in the running buffer was introduced to the double-T crossing by the electrokinetic flow, and then the cell was lysed rapidly under the high electric field. AA in the single rat peritoneal mast cell was electrically migrated to the detection end of separation channel and was detected on the WCNTs/P-PB CFMBE at +0.4 V (vs. Ag/AgCl). The amount of AA in single rat peritoneal mast cells was detected to be 2.4-7.1 fmol. This is the first report of AA in single rat peritoneal mast cells determined by microfluidic chip-ED using MWCNTs/P-PB CFMBE.In the chapter six, A new method of the detection of GSH was developed by the self-assemble microfluidic chip electrochemiluminescence detection system based on tris(2,2′-bipyridine) ruthenium(II) (Ru(bpy)32+). The effects of the concentration of Ru(bpy)32+, running buffer pH, running buffer concentration and separation voltage on microfluidic chip-ECL were studied. The optimum conditions of separation and detection are 1.0×10-4 mol·L-1 for the concentration of Ru(bpy)32+, 3.8×10-3 mol·L-1 NaH2PO4- 1.68×10-2 mol·L-1 Na2HPO4 (pH 7.4) for the buffer solution, 1.2 kV for the separation voltage, 1.0 kV and 5 s for the injection voltage and the injection time, and 1.4 V (vs. Ag/AgCl) for the detection potential. The response for a series of seven injections of 1.0×10-4 mol·L-1 GSH resulted in a relative standard deviation of 0.92% for the migration time, and 2.1% for the ECL intensity, respectively. Experiments showed that this detection system was stable and reliable.
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