Establishment And Characterization Of A Chemiluminescence Microfluidic Sequential Injection Biochemical Analysis System

Microfluidic chip is core technology in miniaturized total analysis systems, which is characterized in portability, high integration, fast speed and low consumption of reagent and sample. Sequential injection analysis system is simple, and sample amount and reagent sequence could be programmed. Chemiluminescence (CL) detection system is also simple, but very sensitive. It become a good detection method for miniaturized total analysis systems. Based on equential injection principle, microfluidic chip and chemiluminescence could be interfaced, and may find good applications for bioanalysis, especially in flow pyrosequencing analysis. Spacial isolation of enzyme and chemiluminescence reactions and independent temperature control of the reactions are important for system optimization.In chapter one, special attention was payed to the introduction of pyrosequencing, and the principle, main influencing factors and application development was reviewed. It is noticed that maintaining the thermal stability of luciferease, proper increase of the extension reaction temperature could increase the realiability of sequencing.In chapter two, a CL microfluidic sequential injection bioanalysis system was established and the performance was investigated taking glucose enzymatic reaction and luminol based CL detection as a model. Firstly, glucose oxidase was immobilized on the Affimag SLE magnetic besds, and the beads were caputured by a magnet to form a flow enzyme microreactor. The Y shape microfluidic chip system was established by capillary, and the enzymatic reaction and chemiluminescent reaction were well separated spacially. Possible application of the flow system for detection of glucose was preliminary demonstrated.In chapter three, a zone thermal control unit was introduced in the flow microfluidic chip system for enzyme reaction temperature optimization. Thermal control was implemented by using ITO glass as the heating element, and platinum resistance as the temperature sensor. The effect of temperature on the enzymatic reaction of glucose was well observed, and the reaction condition was optimized.The flow microfluidic bioanalytical system is easy to establish and control, and may be used to overcome some of the problems in pyrosequencing, such as poor correlation between the signal intensity and the number of repeated bases.