No description available.

No description available.

Abstract As blue/green laser has a short wavelength radiation, it can be applied to micro-machining, laser display, high-density optical data storage, underwater communications, and so on. Large efforts have been devoted to simplify the laser system and reduce the cost. Therefore, various types of blue/green lasers have been studied, especially intracavity frequency doubling of the diode-pumped solid-state laser, which can effectively generate high blue/green laser powers with long lifetime. Among all cavity designs, direct-coated composite crystal is the most compact structure. Compare with the green laser, which is a four-level laser, blue laser belongs to a quasi-three-level system. Thus, it is more important to control the temperature of gain medium. Before the experiment, making an estimation will greatly benefit the progress and efficiency. ZEMAX was utilized to simulate the focus system and GLAD was used to model our intracavity frequency-doubled blue laser. In the experiment, we used a LD array as a pumping source and arranged suitable lens to reduce the array¡¦s spot size. The laser crystal was mounted onto a copper mount which was cooled by the Vapochill cooling system. In addition, we also tried to rotate the crystal and obtained a peak power of 192 mW with only 4 mm cavity length. The result is the highest output power of microchip blue laser to our knowledge. Using lens duct as the pump transport optics can further miniaturize this composite-chip blue laser.

Esta tese de doutorado aborda o desenvolvimento de sistemas microfluídicos eletroquímicos ultrassensíveis mediante a integração de eletrodos i) concêntricos e ii) nanoestruturados seletivos à detecção condutométrica sem contato acoplada capacitivamente (capacitively coupled contactless conductivity detection, C4D) e à amperometria, respectivamente. O uso dos eletrodos concêntricos, uma configuração inédita em microdispositivos na qual o filme fino metálico circunda todo o microcanal, se mostrou efetivo na melhora da detectabilidade da C4D para análises em fluxo de soluções padrão de LiClO4. O limite de detecção (LD) para esse sal foi igual a 343 pmol L-1, um valor aproximadamente quatro ordens de grandeza inferior àquele obtido com eletrodos planares. O microchip amperométrico nanoestruturado, por sua vez, consistiu de filmes de Au modificados com nanotubos de carbono de parede única (single-walled carbon nanotubes, SWCNTs) verticalmente alinhados e foi aplicado a padrões do neurotransmissor serotonina. A melhora na detectabilidade do método foi novamente apreciável; os valores de LD foram de 11,8 (Au liso) e 0,2 nmol L-1 (Au modificado com os SWCNTs verticalmente alinhados). Esse último é menor frente à grande maioria dos valores descritos na literatura, para os quais técnicas diversas foram empregadas, incluindo: i) potenciometria com eletrodos modificados (1,0 a 500 nmol L-1), ii) HPLC-MS (18,2 nmol L-1), iii) eletroforese capilar combinada com etapas de extração, empilhamento e préconcentração do analito (7,9 nmol L-1) e iv) sensor químico (200 nmol L-1). Finalmente, objetivando a fabricação de microchips de vidro, condutométricos e amperométricos, incorporando eletrodos concêntricos nanoestrurados, uma nova técnica de selagem foi desenvolvida. Essa técnica, designada como selagem adesiva de sacrifício, baseia-se no uso do resiste negativo SU-8 como camada intermediária de modo a permitir a vedação entre duas lâminas de vidro. Numa etapa posterior, a remoção seletiva do SU-8 sob o microcanal é realizada. Logo, canais microfluídicos com propriedades de superfície similares às do vidro foram obtidos. O protocolo experimental adotado é i) simples, ii) rápido, iii) não envolve níveis de pressão e temperatura elevados e iv) prescinde o uso de salas \"limpas\". Vedações com forças de adesão satisfatórias foram alcançadas, suportando pressões superiores a 4 MPa. / This PhD thesis reports the development of ultrasensitive electrochemical microfluidic systems by integrating concentric and nanostructured electrodes selective to capacitively coupled contactless conductivity detection (C4D) and amperometry, respectively. The use of the concentric electrodes, a new assembly in microdevices with thin films wrapping around the microchannel, showed to be effective towards improvement of the detectability in pressure-driven flow platforms incorporating C4D. The limit-of-detection (LOD) in flow analysis of LiClO4 solutions was 343 pmol L-1, ca. four orders of magnitude lower than to the levels obtained with planar electrodes alone. The nanostructured amperometric microchip, in turn, is related to integration of vertically aligned singlewalled carbon nanotubes (SWCNTs) over Au film. Such platform was applied to determination of serotonin standards. The nanomaterial influenced remarkably the sensitivity and detectability. Our system achieved a LOD of 0.2 nmol L-1, to the best of our knowledge one of the lowest values reported in the literature. Finally, in order to fabricate glass microdevices, conductometric and amperometric, with nanostructured concentric electrodes, we developed a new bonding method. This technique, called as sacrificial adhesive bonding, is based on SU-8 negative resist like intermediate layer so to allow the sealing between two glass slides. Next, the selective removal of the SU-8 under the microchannel is carried out. Thus, microfluidic channels presenting glass-like surface properties were achieved. The experimental protocol is simple and fast. In addition, neither high-pressure and elevated-temperature nor the use of \"clean\" rooms were not required. Bondings with satisfactory adhesion forces were obtained, supporting pressures above 4 MPa.

Neste trabalho foram desenvolvidas tecnologias e novos processos de fabricação de microdispositivos analíticos visando o acoplamento desta plataforma microfluídica com a detecção condutométrica sem contato (C4D). Uma segunda etapa do trabalho concentrou esforços em melhorar os níveis de detectabilidade da C4D. Para tanto foi proposto um processo rápido e robusto para selagem de canais de vidro para produção de microdispositivos analíticos para eletroforese. Os canais de vidro foram fabricados por processo fotolitográfico e corrosão química em via úmida. Os microcanais obtidos foram selados contra outra lâmina de vidro previamente recoberta por uma membrana polimérica (PDMS) de 50 µm de espessura. Esta mesma membrana, além de promover a junção das placas de vidro, promoveu também o isolamento elétrico entre a solução no interior do canal microfluídico (de uma lâmina de vidro) e os eletrodos metálicos presentes no substrato de vidro oposto. Assim, foi possível acoplar a detecção condutométrica sem contato (C4D) com a plataforma eletroforética proposta. O desempenho analítico desta foi avaliado usando detecção por fluorescência induzida a laser (LIF) e C4D. Eficiência de aproximadamente 47000 pratos/m foi alcançada com boa repetibilidade chip-a-chip. Fluxo eletrosmótico (EOF) estável foi observado apesar da presença do material polimérico compondo parte da parede interna do canal. Com a metodologia proposta, um chip pode ser fabricado em menos de 120 min, já incluindo as etapas de gravação por fotolitografia, corrosão e selagem. Quando comparada à selagem térmica, além do ganho de tempo e facilidade no manuseio dos substratos, o método proposto não necessita de altas temperaturas e os dispositivos obtidos apresentam repetibilidade satisfatória para análises em diferentes dias e em diferentes microchips. A plataforma analítica desenvolvida foi utilizada em um estudo cinético no qual foi possível determinar os parâmetros cinéticos (Vmax = 12,64 mmol L-1 min-1 e KM = 23,8 mmol L-1) da reação de decomposição de ureia catalisada pela enzima urease. Na segunda etapa do trabalho, foi proposta a alteração de um parâmetro físico, a constante dielétrica, da membrana de PDMS usada como isolante de modo a obter um acoplamento capacitivo mais eficiente e como conseqüência uma melhor resposta no detector. Uma discussão teórica fez-se necessária a respeito do princípio de funcionamento da C4D. Os resultados obtidos com experimentos, nos quais a membrana de PDMS foi dopada com dióxido de titânio (TiO2), mostraram que a discussão sobre o funcionamento deste detector está de acordo com as considerações teóricas apresentadas neste trabalho, onde o sinal analítico é proporcional à capacitância e esta é proporcional à constante dielétrica na cela de detecção. Com esta alternativa foi possível reduzir os limites de detecção em experimentos de análise em fluxo de 385,5 para 14,7 µmol L-1 após adição de 50% em massa de TiO2 na membrana de PDMS. / In this thesis were presented technologies developed aiming new manufacturing processes for analytical microdevices by coupling of this microfluidic platform with capacitively coupled contactless conductivity detection (C4D). In a second stage of the work, we focused on improving the levels of detectability of C4D. We proposed a fast and robust process for sealing glass channels to produce analytical microdevices for electrophoresis. The glass channels were fabricated by photolithographic process and chemical wet etching. The obtained microchannels were sealed against another glass plate, which was previously coated with a 50-µm-thick membrane of poly(dimethylsiloxane) (PDMS). The purpose of this membrane, besides promoting the bonding of the two glass plates, was to act as an electrical insulator between the solution within the microfluidic channel on the top glass plate and the metal electrode present on the bottom glass chip. Thus it was possible to couple the contactless conductivity detection (C4D) with the electrophoretic platform proposed. The analytical performance was evaluated using both laser induced fluorescence (LIF) detection and C4D. Efficiency of about 47,000 plates/m was achieved with good chip-to-chip repeatability. Electroosmotic flow (EOF) was observed and stable despite the presence of polymer composing part of the inner wall of the channel. With the proposed methodology, a chip can be manufactured at less than 120 min, including the patterning step by photolithography, chemical etching, and sealing (bonding) step. When compared to the heat sealing procedure, in addition to time savings, and ease of handling of the substrates, the method does not require high temperatures, and the devices obtained show satisfactory repeatability analysis on different days and different microchips. The proposed analytical platform was used in a kinetic study in which it was possible to determine the kinetic parameters (Vmax = 12.64 mmol L-1 min-1 and KM = 23.8 mmol L-1) for the decomposition of urea catalyzed by the enzyme urease. In the second part of this thesis, it was proposed to change a physical parameter, the dielectric constant of the PDMS membrane used as an insulator, to achieve a more efficient capacitive coupling and consequently a better response in the detector. A theoretical discussion was required regarding the operating principle of C4D. The results obtained from experiments in which the PDMS membrane was doped with titanium dioxide (TiO2) showed that the discussion on the functioning of this detector is in agreement with the theoretical considerations presented in this work. The analytical signal was proportional to the capacitance and this was proportional to the dielectric constant in the detection cell. With this alternative we could reduce the detection limits in flow analysis system experiments from 385.5 to 14.7 µmol L-1 after addition of 50% wt of TiO2 in the PDMS membrane.

Esta tese de doutorado aborda o desenvolvimento de sistemas microfluídicos eletroquímicos ultrassensíveis mediante a integração de eletrodos i) concêntricos e ii) nanoestruturados seletivos à detecção condutométrica sem contato acoplada capacitivamente (capacitively coupled contactless conductivity detection, C4D) e à amperometria, respectivamente. O uso dos eletrodos concêntricos, uma configuração inédita em microdispositivos na qual o filme fino metálico circunda todo o microcanal, se mostrou efetivo na melhora da detectabilidade da C4D para análises em fluxo de soluções padrão de LiClO4. O limite de detecção (LD) para esse sal foi igual a 343 pmol L-1, um valor aproximadamente quatro ordens de grandeza inferior àquele obtido com eletrodos planares. O microchip amperométrico nanoestruturado, por sua vez, consistiu de filmes de Au modificados com nanotubos de carbono de parede única (single-walled carbon nanotubes, SWCNTs) verticalmente alinhados e foi aplicado a padrões do neurotransmissor serotonina. A melhora na detectabilidade do método foi novamente apreciável; os valores de LD foram de 11,8 (Au liso) e 0,2 nmol L-1 (Au modificado com os SWCNTs verticalmente alinhados). Esse último é menor frente à grande maioria dos valores descritos na literatura, para os quais técnicas diversas foram empregadas, incluindo: i) potenciometria com eletrodos modificados (1,0 a 500 nmol L-1), ii) HPLC-MS (18,2 nmol L-1), iii) eletroforese capilar combinada com etapas de extração, empilhamento e préconcentração do analito (7,9 nmol L-1) e iv) sensor químico (200 nmol L-1). Finalmente, objetivando a fabricação de microchips de vidro, condutométricos e amperométricos, incorporando eletrodos concêntricos nanoestrurados, uma nova técnica de selagem foi desenvolvida. Essa técnica, designada como selagem adesiva de sacrifício, baseia-se no uso do resiste negativo SU-8 como camada intermediária de modo a permitir a vedação entre duas lâminas de vidro. Numa etapa posterior, a remoção seletiva do SU-8 sob o microcanal é realizada. Logo, canais microfluídicos com propriedades de superfície similares às do vidro foram obtidos. O protocolo experimental adotado é i) simples, ii) rápido, iii) não envolve níveis de pressão e temperatura elevados e iv) prescinde o uso de salas \"limpas\". Vedações com forças de adesão satisfatórias foram alcançadas, suportando pressões superiores a 4 MPa. / This PhD thesis reports the development of ultrasensitive electrochemical microfluidic systems by integrating concentric and nanostructured electrodes selective to capacitively coupled contactless conductivity detection (C4D) and amperometry, respectively. The use of the concentric electrodes, a new assembly in microdevices with thin films wrapping around the microchannel, showed to be effective towards improvement of the detectability in pressure-driven flow platforms incorporating C4D. The limit-of-detection (LOD) in flow analysis of LiClO4 solutions was 343 pmol L-1, ca. four orders of magnitude lower than to the levels obtained with planar electrodes alone. The nanostructured amperometric microchip, in turn, is related to integration of vertically aligned singlewalled carbon nanotubes (SWCNTs) over Au film. Such platform was applied to determination of serotonin standards. The nanomaterial influenced remarkably the sensitivity and detectability. Our system achieved a LOD of 0.2 nmol L-1, to the best of our knowledge one of the lowest values reported in the literature. Finally, in order to fabricate glass microdevices, conductometric and amperometric, with nanostructured concentric electrodes, we developed a new bonding method. This technique, called as sacrificial adhesive bonding, is based on SU-8 negative resist like intermediate layer so to allow the sealing between two glass slides. Next, the selective removal of the SU-8 under the microchannel is carried out. Thus, microfluidic channels presenting glass-like surface properties were achieved. The experimental protocol is simple and fast. In addition, neither high-pressure and elevated-temperature nor the use of \"clean\" rooms were not required. Bondings with satisfactory adhesion forces were obtained, supporting pressures above 4 MPa.

Neste trabalho foram desenvolvidas tecnologias e novos processos de fabricação de microdispositivos analíticos visando o acoplamento desta plataforma microfluídica com a detecção condutométrica sem contato (C4D). Uma segunda etapa do trabalho concentrou esforços em melhorar os níveis de detectabilidade da C4D. Para tanto foi proposto um processo rápido e robusto para selagem de canais de vidro para produção de microdispositivos analíticos para eletroforese. Os canais de vidro foram fabricados por processo fotolitográfico e corrosão química em via úmida. Os microcanais obtidos foram selados contra outra lâmina de vidro previamente recoberta por uma membrana polimérica (PDMS) de 50 µm de espessura. Esta mesma membrana, além de promover a junção das placas de vidro, promoveu também o isolamento elétrico entre a solução no interior do canal microfluídico (de uma lâmina de vidro) e os eletrodos metálicos presentes no substrato de vidro oposto. Assim, foi possível acoplar a detecção condutométrica sem contato (C4D) com a plataforma eletroforética proposta. O desempenho analítico desta foi avaliado usando detecção por fluorescência induzida a laser (LIF) e C4D. Eficiência de aproximadamente 47000 pratos/m foi alcançada com boa repetibilidade chip-a-chip. Fluxo eletrosmótico (EOF) estável foi observado apesar da presença do material polimérico compondo parte da parede interna do canal. Com a metodologia proposta, um chip pode ser fabricado em menos de 120 min, já incluindo as etapas de gravação por fotolitografia, corrosão e selagem. Quando comparada à selagem térmica, além do ganho de tempo e facilidade no manuseio dos substratos, o método proposto não necessita de altas temperaturas e os dispositivos obtidos apresentam repetibilidade satisfatória para análises em diferentes dias e em diferentes microchips. A plataforma analítica desenvolvida foi utilizada em um estudo cinético no qual foi possível determinar os parâmetros cinéticos (Vmax = 12,64 mmol L-1 min-1 e KM = 23,8 mmol L-1) da reação de decomposição de ureia catalisada pela enzima urease. Na segunda etapa do trabalho, foi proposta a alteração de um parâmetro físico, a constante dielétrica, da membrana de PDMS usada como isolante de modo a obter um acoplamento capacitivo mais eficiente e como conseqüência uma melhor resposta no detector. Uma discussão teórica fez-se necessária a respeito do princípio de funcionamento da C4D. Os resultados obtidos com experimentos, nos quais a membrana de PDMS foi dopada com dióxido de titânio (TiO2), mostraram que a discussão sobre o funcionamento deste detector está de acordo com as considerações teóricas apresentadas neste trabalho, onde o sinal analítico é proporcional à capacitância e esta é proporcional à constante dielétrica na cela de detecção. Com esta alternativa foi possível reduzir os limites de detecção em experimentos de análise em fluxo de 385,5 para 14,7 µmol L-1 após adição de 50% em massa de TiO2 na membrana de PDMS. / In this thesis were presented technologies developed aiming new manufacturing processes for analytical microdevices by coupling of this microfluidic platform with capacitively coupled contactless conductivity detection (C4D). In a second stage of the work, we focused on improving the levels of detectability of C4D. We proposed a fast and robust process for sealing glass channels to produce analytical microdevices for electrophoresis. The glass channels were fabricated by photolithographic process and chemical wet etching. The obtained microchannels were sealed against another glass plate, which was previously coated with a 50-µm-thick membrane of poly(dimethylsiloxane) (PDMS). The purpose of this membrane, besides promoting the bonding of the two glass plates, was to act as an electrical insulator between the solution within the microfluidic channel on the top glass plate and the metal electrode present on the bottom glass chip. Thus it was possible to couple the contactless conductivity detection (C4D) with the electrophoretic platform proposed. The analytical performance was evaluated using both laser induced fluorescence (LIF) detection and C4D. Efficiency of about 47,000 plates/m was achieved with good chip-to-chip repeatability. Electroosmotic flow (EOF) was observed and stable despite the presence of polymer composing part of the inner wall of the channel. With the proposed methodology, a chip can be manufactured at less than 120 min, including the patterning step by photolithography, chemical etching, and sealing (bonding) step. When compared to the heat sealing procedure, in addition to time savings, and ease of handling of the substrates, the method does not require high temperatures, and the devices obtained show satisfactory repeatability analysis on different days and different microchips. The proposed analytical platform was used in a kinetic study in which it was possible to determine the kinetic parameters (Vmax = 12.64 mmol L-1 min-1 and KM = 23.8 mmol L-1) for the decomposition of urea catalyzed by the enzyme urease. In the second part of this thesis, it was proposed to change a physical parameter, the dielectric constant of the PDMS membrane used as an insulator, to achieve a more efficient capacitive coupling and consequently a better response in the detector. A theoretical discussion was required regarding the operating principle of C4D. The results obtained from experiments in which the PDMS membrane was doped with titanium dioxide (TiO2) showed that the discussion on the functioning of this detector is in agreement with the theoretical considerations presented in this work. The analytical signal was proportional to the capacitance and this was proportional to the dielectric constant in the detection cell. With this alternative we could reduce the detection limits in flow analysis system experiments from 385.5 to 14.7 µmol L-1 after addition of 50% wt of TiO2 in the PDMS membrane.

The research presented in this work explores the application of microfluidics to the field of proteomics through the design of a multi-channel microfluidic platform and the investigation of individual components of the system. The design of this microfluidic device allows the integration of several protein sample preparation steps for automated electrospray ionization mass spectrometric (ESI-MS) analysis, including protein separation, fractionation and collection, preconcentration and cleanup, and protein digestion. In order for the multi-channel system to function properly, I first evaluated each individual component of the device. Several areas were explored: (i) optimization of polymer monolith for solid-phase extraction (SPE) preconcentration; (ii) investigation of cationic coatings for microchannel surface modification to facilitate positive electrospray of peptides and proteins for chip-MS coupling; (iii) combination of the hydrophobic monolith and the PolyE-323 coating in a single channel device for on-chip SPE and on-bed tryptic digestion with on-line coupling to ESI-MS. Multiplexed microfluidic devices for protein analysis, which integrate a series of microfluidic features, were then designed, built and tested. The multiplexed microfluidic architecture employed a separation channel, a fractionator, an array of microchambers to accommodate monolithic polymer for SPE preconcentration, and an elution channel for the detection of eluted sample using fluorescence detector or mass spectrometer. The performance of the multiplexed devices for integration of multiple analytical steps was explored with sequential fractionation, collection, and elution of fluorescent sample, evaluating the ability to trap and release individual fractions without cross-contamination. Thorough analysis of each of the individual components on the multiplexed microfluidic platform provides valuable insights into the design of such systems, which brings us closer to our final goal of a proteomic processing microchip.

No description available.

Microfabricated thermal cyclers for nucleic acid amplification by using polymerase chain reaction (PCR) have been demonstrated by several groups over the last decade, with improved cycling speed and smaller volumes when compared to conventional bench-top cyclers. However, high fabrication costs coupled with difficulties in temperature sensing and control remain impediments to commercialization. In this study we have used a silicon-glass device that takes advantage of the high thermal conductivity of silicon but at the same time utilizes minimum number of fabrication steps to make it suitable for disposable applications. The thermal cycler is based on noncontact induction heating developed in this group. The microchip reaction kinetics is studied for the first time in-situ during PCR, using a real-time fluorescence block that is capable of data acquisition every 0.7 s from the microchip. The fluorescence information from SYBR green I dye is used to optimize microchip amplification reactions and confirm the product by melting curve analysis. We have also developed a novel non-contact temperature sensing technique using SYBR green fluorescence that can be used for miniaturized PCR devices. The thesis is organized into the following chapters. In chapter 1 we introduce the basic biology ideas that are required to understand DNA amplification. DNA based analysis requires amplification of low initial concentrations to above detectable limits using a technique known as polymerase chain reaction (PCR). In this process, the sample is cycled through three thermal steps for 3040 times to produce multiple copies of DNA. In microchip PCR, conventional polypropylene tubes using 2050 µL volume are replaced by miniaturized devices using ~1 µL sample volumes. The device response improves in terms of ramp rate and total analysis time due to the small volume and smart design of the materials. In this chapter we summarize some of the issues important for miniaturized PCR devices and compare them with commercial tube PCR systems. In chapter 2 we describe the induction heating technique that was developed by our group for miniaturized devices. Induction heating is a noncontact heating technique unlike resistive heating which has been commonly used for microchip PCR. Though resistive heating is very efficient in terms of heat transfer efficiency, it is not suitable for disposable devices and requires multi-step microfabrication. Other non-contact heating techniques such as hot air and IR heating require larger size arrangements that are not suitable for miniaturized devices. The heating was verified by using a thermocouple soldered at the back of the secondary plate that was also used for feedback to the comparator circuit for control. The simple on-off circuit was able to control within ±0.1 ◦C with heating and cooling ramp rates of 25 ◦C/s and 2.5 ◦C/s respectively. In this chapter, we also describe the design and fabrication of the silicon-glass microchip fabricated in our lab. We have used silicon-glass hybrid device for PCR in which glass with a 2 mm drilled hole is anodically bonded to an oxidized silicon surface. The hole formed the static reservoir for 3 µL volume of amplification solution. During PCR, the solution needs to be cycled to high temperature of ~95 ◦C. Hence it was necessary to seal the tiny droplet of liquid against evaporation at this temperature. The devices after being filled by sample were covered by 4 µL of mineral oil to serve as an evaporation barrier. It was easy to recover the whole sample after amplification for further testing. Chapter 3 describes the development of a fluorescent block for SYBR green I dye (SG) used for real-time monitoring of the amplification. The block contains a blue LED for excitation, a dichroic beamsplitter, and silicon photodiode along with filters and focusing optics. Signal levels being weak, we incorporated lock-in detection technique. A TTL at 190 Hz was used to pulse the excitation source and detect the emission at the same frequency using a commercial lock-in amplifier. The block was first characterized using a commercial thermal cycler and polypropylene tubes with different dilution of initial template copy number, and the results crosschecked with agarose gel electrophoresis. Performing continuous monitoring every 0.7s within cycles, we discovered interesting features during extension which have not been studied previously. During the constant temperature extension step, the fluorescence shows a rise and then saturates until the temperature is cycled to the next set point. We have confirmed the same behavior in single cycle extension control experiments and established its connection with polymerase extension activity. We were thus able to extract the activity rate for two different kinds of polymerase in-situ during PCR. By monitoring PCR reactions with different fixed extension times, we were able to determine the optimum conditions for tube PCR. Chapter 4 implements the ideas of fluorescence monitoring from tube that was explained in the previous chapter for the silicon-glass microchip. Since the microchip uses parameters such as sample volume, ramp rates, stay time etc. which are different from tube PCR, we performed several initial test experiments to establish key capabilities such as low volume detection, 3 µL amplification, surface passivation of silicon-glass etc. The same fluorescence block was used to obtain DNA melting point information by continuously monitoring ds-DNA with SG while the temperature is ramped slowly (melting curve analysis). Depending on ds-DNA present, the fluorescence gives a melting temperature (TM ), which was used to calibrate the mix temperature with respect to the thermocouple sensor. After successfully calibrating the microchip, we confirmed complete chip PCR in silicon-glass devices using induction heater. The continuous monitoring of chip PCR gave similar curves as obtained previously for tubes except that the signal level was lower in silicon devices. Extension fluorescence information was used to find an optimum temperature for microchip that shows a maximum activity rate. Similarly the reaction time was optimized in-situ during PCR by using continuous fluorescence data in a feedback experiment. The commercial lock-in amplifier was also replaced by a homemade circuit to successfully pickup fluorescence signal from the microchip during melting curve analysis. In chapter 5, we describe a novel technique to sense the temperature from the microchip without touching the sample volume. Usually the temperature is monitored by a sensor spatially separated from the mix and it has always been challenging to measure the exact temperature accurately. Most of the sensors are not biocompatible and too large in size to be placed inside the small volume of liquid. We have developed a protocol that involves SG fluorescence with addition of excess sensor DNA to the amplification solution. The sensor DNA added into the mix is non specific to the primer used for amplification of the template. It therefore does not participate in the amplification and its number remains unchanged throughout the 3040 cycles of PCR. If the amount of sensor DNA is titrated accurately, it will saturate the fluorescence envelope which then shows very reproducible thermal response with cycling. We have used this thermal response of the fluorescence for feedback as a temperature sensor. The fluorescence feedback was shown to produce identical amount of product in comparison to thermocouple feedback. The product can also be verified by melting curve analysis if the sensor DNA is chosen carefully depending on the product. In this chapter we also discuss some preliminary experiments with smart devices that will use dye based temperature sensor and control along with fluorescence based amplification monitoring. Chapter 6 summarizes the thesis and discusses some of the future areas which can be explored in the field of microchip PCR devices.