Photopatterning And Applications Of Poly(N-isopropylacrylamide) In Microfluidic Chip

Microfluidic chips,which possess a number of useful capabilities,the ability to use very small quantities of samples and reagents,low cost,short times for analysis,integration for analytical devices,and small size for the control of concentrations of molecules in space and time,have found widely conventional applications in screen conditions for assays,separations coupled to detection device,and synthesis of organic or inorganic compounds etc.Currently,with the rapidly increasing demands for high-throughput,automated,integrated microfluidic system,the conventional microfluidic technology shows too many shortcomings in these complex assays which include multiple experimental steps,different functional regions,and various fluids,so the design and fabrication of functional microstructures in microchips have attracted substantial interest and become more and more important for bioanalysis,clinical diagnostics,drug screening and delivery,tissue engineering,cell culture,and life science.Thus,the development of spatially separate regions with different properties is faced with extreme challenge.Stimuli-responsive polymers,known as "smart materials",have many significant advantages over others owing to their ability to undergo adrupt volume,surface structure and property changes in response to the surrounding environment without the requirement of an external power source.Poly(N-isopropylacrylamide)can be taken as a representative of a class of temperature-sensitive polymers,it exhibits not only volume transition but also surface wettability changes at around 32 ?(lower critical solution temperature,LCST).Combined with spatially controlled photopatteming technology,it is simple and convenient to create these microstructures with different functions at any locations of microchannels.Over the past decade,numerous publications have been emerged for the applications of patterning approach in the area of microfluidic chip.In this dissertation,we have designed three photopattemed microstructures for analytical devices.1.Fabrication of microporous stimuli-responsive poly(N-isopropylacrylamide)monolith and its applications in glucose microfluidic biosensorsWe demonstrated a new strategy for replaceable enzymatic microreactor based on a switchable wettability interface of poly(N-isopropylacrylamide)(PNIPAAm).PNIPAAm porous polymer monolith(PPM)with 3D macroporous framework is photopolymerized in glass microchip within 30 seconds.The PNIPAAm PPM not only shows its reversible swelling/shrinking property at the different temperature around the lower critical solution temperature(LCST),but also shows reversible hydrophilicity/hydrophobicity corresponding to its swelling/shrinking status.Based on these properties,a biocompatible and replaceable on-chip enzymatic microreactor has been successfully built by means of the reversible adsorption and release of glucose oxidase(GOx)on the robust and stable matrix.Coupled with a carbon fiber microelectrode as electrochemical detector,the microreactor has been successfully employed for the detection of glucose in human serum samples.This approach may provide a promising way for high efficient and renewable microreactors that will find wide application in clinical diagnosis,biochemical synthesis/analysis,and proteomic research.2.Fabrication of nanoporous poly(N-isopropylacrylamide)membrane and its applications in DNA enrichment and releaseAn ideal nanoporous polymer membrane(poly(N-isopropylacrylamide),PNIPAAm)with uniform,dense,robust and ion-permeable structure has been successfully fabricated in glass microchannels by means of in situ spatially controlled sophisticated photopatterning technology.It possesses the greatly improved performances in enrichment factor and release ability for biomolecules such as DNA after deliberately optimizing the experimental parameters and conditions including the compositions of precursor,polymerization conditions,buffer systems,membrane lengths.Over 5000 enrichment factor of the nanoporous polymer membrane for DNA stacking followed by a complete release in elution step is successfully achieved.In addition,the concentration polarization phenomena have been alleviated by the ingenious chip design of continuous gravitational flow on the both sides of the membrane.It is anticipated that such a membrane could be potentially applied for low-abundant analytes detection in the assay,especially for nascent diagnosis of trace cancer biomarkers,where the high sensitively is strongly required.3.Construction of an integrated preconcentration microchip and its applications in aptamer affinity probe gel electrophoresis assaysBased on the predictable behavior of nucleic acid aptamer in electrophoresis as a result of uniform charge-to-size ratios with low molecular weight and high negative charge,it would be benefit for the affinity capillary electrophoresis,demonstrating a fully integrated microchip plateform toward portable point-of-care analysis.So we describe here a aptamer-based affinity capillary electrophoresis assay of thrombin binding system in an integrated microfluidic chip,it includes two functions,a nanoporous membrane element for preconcentration and a long gel sieving matrix for separation.The key concept is constructing a versatile plateform to improve sensitivity of detection and simultaneously achieve efficient separation of the complex from its aptamer.The spatially controlled photopatterning technology is used to create poly(N-isopropylacrylamide)membrane microstructure inside the sepcial location of glass microchip,and the methylcellulose(MC)gel solutions are loaded adjacent to the membrane in the separation channel enabled zero dead volume integration of both functions.The confocal laser-induced fluorescence detection system(LIF)provides quantitation of protein targets.In order to evaluate the performance of integrated microchip,a more simple double-T microchip is designed to be a reference without preconcentration,significant signal enhancement and good peak resolution of thrombin complex is achieved.The method has broad applicability to improves sensitivities of various other affinity shift electrophoretic assay.