| There are a great of various chemical substances in the environmental pollutants. Due to characteristics of interaction and trace level content, it promotes the development of miniature bioassay instrument with automation based on the new method and technology, which can meet the demand of detection techniques with advantages of high sensitivity, high selectivity, in situ, in-vivo, and field quick analysis in environmental science. The SPR sensor associated with immunoassay can monitor biomolecular interaction on-line, including determinations of reactant type and analyte concentration without label, separation and purification. With the virtues of high detection sensitivity, fast response, universality and anti-electromagnetism interference, it offers advantages when be applied to environmental pollutants detection.To satisfy the needs of environmental pollutants detection, associated with the flow injection analysis technique, a miniature SPR bio-analytical instrument with automation was developed, whose kernel is SpreetaTM sensor by TI. After optimization of the instrument performance, it acted as the detector of HPLC. Finally, associated with immunoassay, the SPR biosensors were fabricated to detect the trace amount algae toxins in the surface water and seawater.The main research achievements include the followings.(1) According to the Maxwell equations and its boundary condition that the electromagnetic wave propagates in the medium, and the Drude model of free-electron gas in the mental, the principle that surface plasma resonance was excited by the incident light was interpreted in detail. Based on the theory of attenuated total reflection of light, the reflectance equation of Kretschmann SPR sensor was derivated. The performance of the senor influenced by each component was simulated. The complex film with the structure of'prism-Ag film-Au film'in SPR sensor was designed. Its parameters were optimized via simulation. It could be concluded that the SPR sensor with 50nm complex film, which includes 20nm gold film, provides the best performance.(2) Based on the Spreeta sensor TSPR1A170100 by TI Ltd, the flow injection analysis technique and the design method of virtual instruments, the miniature SPR bio-analytical instrument with automation was developed. It can perform the bioassay function of clean, injection and regeneration, and detect 8 different samples in sequence. Coupled with immunoassay method, it can be well satisfy the needs of accurate, rapid, and field detection for environmental pollutions.(3) The performance of the fabricated SPR instrument was optimized. After the temperature effect of the instrument was studied, the efficiently constant temperature regulator was developed to improve the sensor's performance. Based on the efficiently constant temperature regulator, the variation of analyte's refractive index against temperature was evaluated. Therefore, the temperature compensation method for the refraction index of the liquid determined via SPR sensor was proposed firstly. The compensated result was two degree lower than the refraction index without compensation determined by SPR sensor. To overcome the defect that the SPR formant curve interfered by the low-frequency noise, which was difficult to be avoided by traditional analysis algorithm for SPR formant curve, the HHT digital filter for the non-stationary nonlinear signal was designed and firstly applied to filter the continuous monitoring signal of SPR. It can help to evidently improve the signal noise ratio, and the detection limit was 8 lower than that without filtering.(4) Based on the SPR bio-analytical instrument, its performance is evaluated via experiment. Since the SPR sensor is sensitive to the changes of the refractive index of the solution, it can be applied to detect concentration of the defined composition in solution. It was proposed that the SPR acted as the novel detector of the flow analysis system. The feasibility was confirmed by the experiment that SPR detector was coupled to HPLC. Result showed that it fit for the flow analysis system due to its universality, low sample consumption and cost-effectiveness. The SPR detector is of advantage to detecting the composition insensitive to ultraviolet detection or fluorescence detection, or the composition needed to be derived for determination, such as glucide, lipid, high polymer and surfactant.(5) Coupled with the immunoassay method, the miniature SPR bio-analytical instrument with MC-LR biosensor was applied to detection the MC-LR in surface water. The performance was evaluated, and real sample of surface water was determined. Equipped with the OA biosensor, it was firstly applied to detect OA, which belong to the PSP algae in the sea. The method can be universally established to detection the other algae in the sea.The result of the research indicates that miniature SPR bio-analytical instrument with automation can be well applied to detect trace environmental pollution directly and rapidly without cumbersome sample pretreatment due to specificity of immunosensor. In addition, the reusable immunosensor with free-labeling contributes to reducing the cost of assay. All of its characteristics can complement the shortcomings of traditional assay method, such as long assay time, high cost and labeling. Most of all, it is a promising project to monitor field samples in situ for environmental pollution detection.
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