Doctor of Philosophy / Department of Chemistry / Christopher T. Culbertson / Recent approaches in analytical separations are being advanced towards the “lab-on-a-chip” concept in which multiple lab functions are integrated into micro/nano fluidic platforms. Among the variety of separation techniques that can be implemented on microfluidic devices, capillary electrophoresis is the most popular as it provides high efficiency, simple, fast and low cost separations. In addition, integrating miniaturized fluid manipulation tools into microfluidic devices with separations is essential for a variety of biological applications. Chapter 1 discusses the fundamentals of capillary electrophoresis and miniaturized fluid manipulation tools and provides an over view of single cell analysis in microfluidics. In chapter 2, the integration of miniaturized peristaltic pumps into multilayer microfluidic platforms is discussed. In addition, device characterization, precise fluid control and high throughput single cell analysis are discussed. As a proof of principle, T-lymphocytes were loaded with two fluorescent probes Carboxyfluorescein diacetate (CFDA) and Oregon green (OG). Thousands of single cells were automatically transported, lysed on these devices and analytes from the lysate were electrophoretically separated. 1120 cells were analyzed over the course of 80 min (14 cells/min) and separation characteristics of analytes released from individual cells were investigated. In the third chapter, the development of microfluidic platforms for the electrochemical detection of nitric oxide (NO) and other reactive nitrogen species (RNS) at the single cell level is discussed. A microfluidic system was developed to perform rapid cell lysis followed by electrochemical detection. Miniaturized microband electrodes were designed and integrated with a microfluidic separation channel. Three alignment techniques (in-channel, end-channel and off-channel configurations) were used to detect the electrochemical response of the analyte of interest. Furthermore, a model analyte (CFDA) was used to demonstrate the potential of performing the simultaneous dual detection with electrochemical and laser induced fluorescence detection. In addition, the same microfluidic platform was adapted to detect intracellular superoxide using laser induced fluorescence. In the fourth chapter, the off-chip integration of optical fiber bridges with multilayer microfluidic chips is discussed. A multimode optical fiber (~10cm long) was integrated between the single cell lysing spot and a spot downstream of the separation channel in order to detect both intact cells and the analyte in the lysate. This technique was used to create two detection spots on the microfluidic platform with the use of a single excitation source and single detector. Fluorescently labeled T-lymphocytes were automatically transported and lysed in a manner similar to that described in chapter 2. Hundreds of single cells were analyzed and the absolute migration time was determined for the analytes in the lysate. In addition, the separation characteristics of fluorescently labeled protein kinase B peptide substrates were investigated. Furthermore, this technique was used to measure cell size and the velocity of intact cells (discussed in 5th chapter) by making use of a light tunneling concept available in multimode optical fibers. All the experiments presented in this dissertation exploit the use of multilayer microfluidic platforms to investigate intracellular components in single cells in a high throughput manner that has several advantages over current conventional techniques.

Orientador: Nelson Ramos Stradiotto / Resumo: O comportamento cíclico voltamétrico do acetaldeído e seu derivado foi estudado em eletrodo de gota pendente de mercúrio e carbono vítreo. Este estudo foi utilizado na otimização das condições para a detecção eletroquímica desses compostos por Cromatografia Líquida de Alta Eficiência (C.L.A.E.). O acetaldeído foi derivado com 2,4-dinitrofenilhidrazina (DNPHi) e o produto, a 2,4-dinitrofenilhidrazona do acetaldeído (AcH-DNPHo) foi eluida e separada por coluna de fase reversa C18 em condições isocráticas com fase móvel contendo uma mistura binária de metanol-solução aquosa de cloreto de lítio (LiCl) com concentração de 1,0 ' 10-3 M (80:20% v/v) e vazão de 1,0 mL min-1. A detecção eletroquímica (D.E.) do AcH-DNPHo foi realizada por C.L.A.E. com potencial fixado em + 1,0 V vs eletrodo de referência Ag/AgCl. O método proposto foi simples, rápido (tempo de análise 7 min.), com limite de detecção 3,80 mg L-1, altamente seletivo, e reprodutível [desvio padrão relativo 8,2% (n=12)]. A curva analítica do AcH-DNPHo foi linear na faixa compreendida de 0 - 300 mg L-1, com índice de correlação linear 0,9998. A concentração de acetaldeído determinada em amostras de álcool combustível foi de 83 - 341 mg L-1 com desvio padrão de 1 - 6 %, na variação da área de pico e recuperação superior a 99%. A quantidade do teor de acetaldeído encontrado nestas amostras foi significativamente alta e os resultados obtidos foram comparados com o método de detecção espectrofotométrica (UV/Vis) / Abstract: The cyclic voltammetric behaviour of acetaldehyde and their derivatives has been studied at a hanging drop mercury and glassy carbon electrode. This study was used to optimise the conditions for the electrochemical detection of these compounds following high-performance liquid chromatographic (H.P.L.C.) separation. The acetaldehyde was derivatized with 2,4-dinitrophenylhydrazine (DNPHi) and the product 2,4-dinitrophenylhydrazone (AcH-DNPHo) was eluted and separated by reversed-phase column C18 in isocratic conditions with mobile phase containing binary mixture of methanol-chloride littium aqueous solution with concentration 1,0 ' 10-3 M (80:20% v/v) and flow rate of 1.0 mL min-1. The electrochemical detection (E.D.) of AcH-DNPHo was performed by H.P.L.C. set at +1.0 V vs Ag/AgCl as reference electrode. The proposed method was simple, rapid (analysis time 7 min), with detection limit 3.80 mg L-1, highly selective, and reproducible [relative standard deviation 8.2 % (n=12)]. The calibration graph for AcH-DNPHo was linear in the range of 0 - 300 mg L-1 with correlation coefficient 0,9998. The concentrations of acetaldehyde determined in fuel alcohol samples was in the range of 83 - 341 mg L-1 with standard deviation of 1 - 6 %, in peak area variation and analytical recovery was >99%. The amount of acetaldehyde content found in these samples was significantly higher and the results obtained were compared to the spectrofotometric detection method (UV/Vis) / Mestre

Submitted by Cláudia Bueno (claudiamoura18@gmail.com) on 2016-02-17T14:00:34Z No. of bitstreams: 2 Dissertação - Gerson Francisco Duarte Júnior - 2015.pdf: 3229718 bytes, checksum: 81bfec5942eb1f9b2ce2a8218a18cdba (MD5) license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) / Approved for entry into archive by Luciana Ferreira (lucgeral@gmail.com) on 2016-02-18T11:48:18Z (GMT) No. of bitstreams: 2 Dissertação - Gerson Francisco Duarte Júnior - 2015.pdf: 3229718 bytes, checksum: 81bfec5942eb1f9b2ce2a8218a18cdba (MD5) license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) / Made available in DSpace on 2016-02-18T11:48:18Z (GMT). No. of bitstreams: 2 Dissertação - Gerson Francisco Duarte Júnior - 2015.pdf: 3229718 bytes, checksum: 81bfec5942eb1f9b2ce2a8218a18cdba (MD5) license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) Previous issue date: 2015-07-06 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / The current work describes some instrumental advances on contactless conductivity detection (C4D) as well as the development of ionic electrodes to be coupled on microchips electrophoresis (MSE). Firstly, was constructed a high-resolution C4D detector based on openC4D electronic. Frequency and amplitude was optimized resulting in 400 kHz and 4 Vpp, respectively, for optimal values. These conditions was able to be used in different MSE devices including polymethylmethacrylate, polydimethysiloxane and polyester-toner. For liquid electrodes fabrication, channels were engraved on a polymeric plate using a CO2 laser system and sealed with adhesive membrane. After that, PDMS microchip was reversibly sealed against the polymer membrane. The electrode channels were filled with different ionc solutions (KCl, NaCl and LiCl). The KCl solution with 2 mol/L provide the best peak intensity. Then analytical performance of the ionic electrode was test in a separation of K+, Na+ e Li+ (300 μmo/L each). A good separation was observed and limit of detection achieved were 28, 40 and 58 μmol/L for K+, Na+ e Li+, respectively. The biggest advantage the use of ionic electrode are related of easiness fabrication process, simplicity and lower cost per device. Additionally, a commercial system of MSE-C4D was propose in a separation of sugars. The analytical separation was realized under the alkaline condition (pH>12), in order to provide the negative charge for sugars. After optimization of buffer and experimental conditions, a mixture containing fructose, galactose, glucose, lactose and sucrose was separated in less 3 min. / O presente trabalho descreve alguns avanços instrumentais relacionados à construção de um detector condutométrico de alta resolução e também ao uso de eletrodos iônicos para detecção condutométrica sem contato (C4D) acoplada aos microssistemas eletroforéticos (MSE). Além disso, o trabalho também descreve o desenvolvimento de metodologia para a análise de açúcares voltadas à aplicações na produção do etanol de 2ª geração. Inicialmente, foi construído e otimizado um detector condutométrico para MSE baseado na eletrônica OpenC4D. Os valores de frequência e amplitude obtidos com a otimização foram de 400 kHz e 4 Vpp, respectivamente. Com os valores otimizados, o detector se mostrou versátil ao utilizá-lo em MSE fabricados em diferentes substratos. Para a construção dos eletrodos iônicos, primeiramente foram gravados canais em placas poliméricas e selados com uma membrana adesiva. Um microchip de PDMS foi selado reversivelmente contra este suporte. Em comparação aos eletrodos metálicos, os eletrodos iônicos oferecem algumas vantagens, tais como, simplicidade, facilidade de fabricação, e baixo custo por dispositivo. Diferentes soluções foram avaliadas como eletrodos, dentre as quais a solução de KCl 2 mol/L forneceu os melhores resultados em termos de intensidade de sinal. O desempenho analítico dos novos eletrodos foi avaliado para a separação de uma mistura padrão de K+, Na+ e Li+ (300 μmol/L cada). Neste estudo, foram obtidos limites de detecção entre 28 e 58 μmol/L. Estes valores são comparáveis aos obtidos com os convencionais eletrodos metálicos. Uma metodologia para separação de frutose, glicose e sacarose utilizando um sistema comercial de MSE-C4D foi proposta. As análises foram baseadas na utilização de um pH alcalino (>12) que garante a ionização destas espécies para a forma aniônica. Após a otimização das condições experimentais onde um tampão composto por NaOH/Na2HPO4 75/10 mmol/\L foi o que forneceu melhores resultados em temos de resolução entre par frutose e glicose. com estas condições uma mistura contendo frutose, galactose, glicose, lactose e sacarose foi separada em um tempo de 3 min.

<p>This thesis ties some electrochemical aspects of development and fabrication of an analytical system on a microchip together. These aspects develop through the fundamentals of amperometric detection in microsystems and microfabrication via the interaction of electrochemical detection and electrophoretic separation finally to the interfacing of a microsystem to the macro world.</p><p>Paper <b>I</b> deals with amperometric detection in microscale systems and describes the fabrication of the necessary on-chip microelectrodes together with fluidic channels in silicon. It was furthermore studied, if the interelectrode distance of some μm could be used to improve the sensitivity in amperometric detection by employing redox cycling. </p><p>Papers <b>II</b>, <b>III</b> and <b>IV</b> deal with the effect of a high voltage field on amperometric detection. In analytical microdevices typically an electrophoretic separation step (e.g. capillary electrophoresis, CE) precedes the detection. The interference of the CE high voltage with the amperometric detection potential is often considered one of the main hindrances for an effective combination of these techniques. In paper <b>II</b> one reason for the observed disturbing potential shift was elucidated. It was shown that positioning of working electrode and reference electrode on an equipotiental surface eliminates this problem. Paper <b>III</b> reports an application of this technique. In paper <b>IV</b> it could be shown that this approach could further be used to significantly reduce the instrumental requirements for amperometric detection in CE.</p><p>Papers <b>V</b>, <b>VI</b>, <b>VII</b>, finally discuss the interfacing of low volumetric flows that typically occur on microanalytical devices to other techniques. Both, interfacing from liquid to liquid phase (μLC to CE in paper <b>V</b>) and from liquid to gas phase (CE to MS in paper <b>VI</b> and <b>VII</b>) were discussed. Electrochemical methods are used in this context to evaluate the stability and, in paper <b>VI</b> and <b>VII</b>, to increase the understanding of underlying processes of corrosion.</p>

Electrochemical detection of metals can be done at polycrystalline gold ultramicroelectrodes using repetitive cyclic voltammetry (RCV), a detection method sharing some similarities with anodic stripping voltammetry (ASV). Each cycle of the potential waveform for RCV involves application of a negative preconcentration potential (for 50 to 300 ms) followed by a cyclic voltammetry (CV) scan at 20 to 1000 V/s. The response due to the metals is evident at potentials negative of the region for oxide formation in the resulting CVs. Metals are deposited at the Au surface by underpotential deposition (UPD) processes. Any metal that can be analyzed by RCV could potentially be quantified using UPD-ASV at Au (rather than by ASV at Hg). The UPD kinetics of Pb and Cu at polycrystalline Au were examined by setting kinetic parameters (rate constant, symmetry factor, and electrosorption valency) within a simulation program used to generate simulated CVs. Reasonably good agreement between experimental and simulated CVs was possible using the simulation, with the same kinetic parameters used to generate simulated CVs to match experimental CVs over a range of sweep rates for each system. Using this method, the following rate constants (k) were estimated: for UPD of Cu in H2SO4 and HClO4, ks ~ 36000 s−1 and 11000 s−1, respectively, and for UPD of Pb in H2SO4, ks ~ 400000 s−1. <p> Repetitive cyclic voltammetry was applied to the detection of metals separated by capillary electrophoresis. Separation of Tl+, Cd2+, Cu2+, Pb2+, Zn2+, Ni2+, Co2+ and Mn2+ was demonstrated in 0.01 mol/L acetic acid and 0.01 mol/L ammonium acetate(pH ~ 4.6) using RCV. While stacking is commonly exploited for sensitivity enhancement during injection, it was shown that detection-end stacking is also useful. A novel technique named electrophoretic extraction (EE) was developed for analysis of particle-containing solutions (e.g. soil extracts or other colloidal suspensions). EE involves application of backpressure during CE to prevent particles from entering the separation capillary: the applied pressure is regulated so analyte ions enter the capillary and migrate to the detector, whereas other particles are prevented from entering the capillary. The feasibility of this approach was demonstrated.

In nanopore analysis, peptides and proteins can be detected by the change in current when single molecules interact with an α-hemolysin pore embedded in a lipid membrane. Studies into the effects of fluorenylmethoxycarbonyl (Fmoc), acetylation or proline modification to negatively charged α-helical peptides showed that Fmoc peptides give more translocations than acetylated peptides. The addition of a proline in the middle of an acetylated peptide further reduces the number of translocations compared to Fmoc. The effect of peptide conformation on translocation or intercalation was studied with small α-helical and β-sheet hairpins. The capped β-hairpin increased translocations compared to the uncapped. The Fmoc-α-helical hairpin, containing a disulfide link, displayed both bumping and translocations whereas in the unlinked peptide the proportion of translocations was greater. Prion diseases arise from the misfolding and aggregation of the normal cellular prion protein. Nanopore analysis of prion peptides with α-helical and β-strand sequences show changes to the event parameters that help distinguish them. The interaction of bovine prion protein (bPrP), with α-hemolysin showed both bumping (type-I) and intercalation/translocation (type-II) events. There are several lines of evidence that indicate these type-II events with a blockade current of -65 pA for bPrP, represent translocations. Nanopore analysis showed that about 37% events were translocations. The interaction of metal ions with bPrP showed that Cu(II) or Zn(II) reduced translocations. Surprisingly, Mn(II) caused an increase in translocation events to about 64%. Complex formation between antibodies and prion peptides and proteins can be detected by nanopore analysis. The PrP/antibody complex is too large to translocate whereas the event parameters for unbound molecules are unchanged. In principle, a nanopore can detect a single molecule; thus, this work represents the first step towards the development of a prion detector. The nanopore will provide the sensitivity and the antibodies will provide the specificity to distinguish between PrPC and PrPSc. Also, the prion N- and C-terminal signal peptides interact with bPrP changing the event parameters, relating to a new mechanism. Finally, the folding intermediates of bPrP at 0.86 M Gdn-HCl suggests that the protein unfolds and then refolds into a different conformation with event parameters similar to those of bPrP.

Methicillin Resistant Staphylococcus aureus (MRSA) is one of the most important threats of nosocomial infections in many regions of the world and Vancomycin Resistant Enterococcus (VRE) is an emerging pathogen that develops full resistance against third-generation glycopeptide antibiotics. Conventional methods for identification of MRSA and VRE generally depend on culturing, which requires incubation of biological samples at least 24-72 hours to get accurate results. These methods are time consuming and necessitate optical devices and experts for evaluation of the results. On the other hand, early diagnosis and initiation of appropriate treatment are necessary to decrease morbidity and mortality rates. Thus, new diagnostic systems are essential for rapid and accurate detection of biological analytes at the point of care. This study presents design, fabrication, and implementation of MEMS based micro electrochemical sensor (&micro / ECS) to detect the methicillin resistance in Staphylococcus aureus and vancomycin resistance in Enterococcus species. To the best of our knowledge, the developed sensor is the first &micro / ECS which utilizes on-chip reference (Ag), working (Au), and counter (Pt) electrodes together with a microchannel to detect MRSA and VRE. The characterization of the designed sensor was achieved analyzing the interactions of the buffer solutions and solvents with the electrodes and Parylene C film layer by using optical and electrochemical methods. Specific parts of genes that are indicators of antimicrobial resistances were used in order to detect the resistances with high selectivity and sensitivity. Thus, synthetic DNA and bacterial PCR product were used as target probes in redox marker based detection and enzyme based detection, respectively. In order to enhance the hybridization, folding structures of the capture probe were investigated by using mfold Web Server. In redox marker based detection, the hybridization of DNA was indirectly detected by using Hoechst 33258 as redox marker with differential pulse voltammetry. The cross reactivity of the tests were performed by using different target probes of femA genes of S. aureus and S. epidermis, which are the major genes detected in methicillin detection assays. Consequently, amplification of signal by using horseradish peroxidase and TMB/H2O2 as substrate was achieved in order to enhance detection sensitivity. The sensor could detect 0.01 nM 23-mer specific part of mecA gene with redox marker based detection and 10 times diluted PCR product with enzyme-based detection in about six hours including the steps of sample preparation from whole blood. This sensor with its compatibility to MEMS fabrication processes and IC technology has a promising potential for a hand-held device for POC through the integration of micropotentiostat.

In nanopore analysis, peptides and proteins can be detected by the change in current when single molecules interact with an α-hemolysin pore embedded in a lipid membrane. Studies into the effects of fluorenylmethoxycarbonyl (Fmoc), acetylation or proline modification to negatively charged α-helical peptides showed that Fmoc peptides give more translocations than acetylated peptides. The addition of a proline in the middle of an acetylated peptide further reduces the number of translocations compared to Fmoc. The effect of peptide conformation on translocation or intercalation was studied with small α-helical and β-sheet hairpins. The capped β-hairpin increased translocations compared to the uncapped. The Fmoc-α-helical hairpin, containing a disulfide link, displayed both bumping and translocations whereas in the unlinked peptide the proportion of translocations was greater. Prion diseases arise from the misfolding and aggregation of the normal cellular prion protein. Nanopore analysis of prion peptides with α-helical and β-strand sequences show changes to the event parameters that help distinguish them. The interaction of bovine prion protein (bPrP), with α-hemolysin showed both bumping (type-I) and intercalation/translocation (type-II) events. There are several lines of evidence that indicate these type-II events with a blockade current of -65 pA for bPrP, represent translocations. Nanopore analysis showed that about 37% events were translocations. The interaction of metal ions with bPrP showed that Cu(II) or Zn(II) reduced translocations. Surprisingly, Mn(II) caused an increase in translocation events to about 64%. Complex formation between antibodies and prion peptides and proteins can be detected by nanopore analysis. The PrP/antibody complex is too large to translocate whereas the event parameters for unbound molecules are unchanged. In principle, a nanopore can detect a single molecule; thus, this work represents the first step towards the development of a prion detector. The nanopore will provide the sensitivity and the antibodies will provide the specificity to distinguish between PrPC and PrPSc. Also, the prion N- and C-terminal signal peptides interact with bPrP changing the event parameters, relating to a new mechanism. Finally, the folding intermediates of bPrP at 0.86 M Gdn-HCl suggests that the protein unfolds and then refolds into a different conformation with event parameters similar to those of bPrP.

Electrochemical detection of metals can be done at polycrystalline gold ultramicroelectrodes using repetitive cyclic voltammetry (RCV), a detection method sharing some similarities with anodic stripping voltammetry (ASV). Each cycle of the potential waveform for RCV involves application of a negative preconcentration potential (for 50 to 300 ms) followed by a cyclic voltammetry (CV) scan at 20 to 1000 V/s. The response due to the metals is evident at potentials negative of the region for oxide formation in the resulting CVs. Metals are deposited at the Au surface by underpotential deposition (UPD) processes. Any metal that can be analyzed by RCV could potentially be quantified using UPD-ASV at Au (rather than by ASV at Hg). The UPD kinetics of Pb and Cu at polycrystalline Au were examined by setting kinetic parameters (rate constant, symmetry factor, and electrosorption valency) within a simulation program used to generate simulated CVs. Reasonably good agreement between experimental and simulated CVs was possible using the simulation, with the same kinetic parameters used to generate simulated CVs to match experimental CVs over a range of sweep rates for each system. Using this method, the following rate constants (k) were estimated: for UPD of Cu in H2SO4 and HClO4, ks ~ 36000 s−1 and 11000 s−1, respectively, and for UPD of Pb in H2SO4, ks ~ 400000 s−1. <p> Repetitive cyclic voltammetry was applied to the detection of metals separated by capillary electrophoresis. Separation of Tl+, Cd2+, Cu2+, Pb2+, Zn2+, Ni2+, Co2+ and Mn2+ was demonstrated in 0.01 mol/L acetic acid and 0.01 mol/L ammonium acetate(pH ~ 4.6) using RCV. While stacking is commonly exploited for sensitivity enhancement during injection, it was shown that detection-end stacking is also useful. A novel technique named electrophoretic extraction (EE) was developed for analysis of particle-containing solutions (e.g. soil extracts or other colloidal suspensions). EE involves application of backpressure during CE to prevent particles from entering the separation capillary: the applied pressure is regulated so analyte ions enter the capillary and migrate to the detector, whereas other particles are prevented from entering the capillary. The feasibility of this approach was demonstrated.

Capillary electrophoresis chip has gradually ripe along with the development of MEMS technology. However, such these electrophoresis chips was design closed-channel form whose process including the micro-channel forming and chip bonding and so on, so the cost is higher. In addition, if these chips use repeated will cause some pollution problems such as obstruction or difficult to clean in the closed-channel. Therefore, to fabricate a non-closed microfluidic chip system will resolve the issues above listed. In this study has successful developed a convenient and low-cost thread microfluidic system, the thin polyester thread is instead of the traditional closed separation channel. And to avoid the cross contaminations that the separation channel can free replace a new electrophoresis separation channel by the roller equipment. Thread microfluidic systems can take advantage of capillary action to move, as a disposable chromatography flow channel, and the electric field is applied to this system for electrophoresis separation and electrochemical detection in the backend. This research develop an novel process technology, the hot embossing technology shape from concave embossing and metal coating procedure in PMMA, the salient pillow-electrode structure has be produced in PMMA board, the salient electrode structure set up the polyester fiber thread, the polyester fiber thread is as the electrophoresis separation channel, and electrochemical detect samples in back-end. In this study take plasma treatment to improve wettability and surface roughness of the polyester fiber thread, in order to improve the operational effectiveness of the thread microfluidic systems. The cyclic voltammetry measure potassium ferricyanide samples and the results showed that the performance of thread microfluidic system significantly increase after the plasma treatment, the measuring current value is 10 times greater than without the plasma treatment, and the estimated detection limit of potassium ferricyanide is around 6.25 £gM in the plasma treatment one. In addition, the thread microfluidic devices with plasma treatment has successful separation and detection the mixing samples of 0.3 mM chlorine, bromine and iodine ions, and the signal of the S/N ratio is 6 times higher than the without plasma treatment one, and the number of theoretical plates of electrophoresis separation also enhance to 28% in the plasma treatment one. In addition, in order to further enhance the detection sensitivity of the thread microfluidic systems, the study design and fabricate the concave-shaped three-dimensional electrode structure, and to achieve greater contact area between thread and the electrode. Thread microfluidic system with plasma treatment detect in concave-shaped three-dimensional electrode and flat electrode respectively, the results showed that concave-shaped electrodes in the potassium ferricyanide detection limit that measured current value is 10 times greater than flat electrode, and a mixture of dopamine and catechol sample are electrophoresis separated that concave-shaped 3D electrode whose measured S/N ratio is 5 times higher than flat electrode, and the number of theoretical plates is 1.5 times higher than the flat electrode. This study develops a novel thread microfluidic system that will provide a simple process and low-cost, and can solve the shortcomings of conventional closed-channel electrophoresis chips. In future, in this study develop the thread microfluidic system architecture will have contribute for fast electrophoresis separation and detection.