| Surface plasmon resonance (SPR) represents one of the rapidly advancing technologies for fast, real-time and in situ detection of chemical and biological analytes. The SPR technique is sensitive to the change of refractive index or thickness of the test medium in vicinity of the metal surface, especially for the molecules with larger mass change. However, the inability of conventional SPR to measure extremely small changes in refractive index impedes its further application in ultrasensitive detection. Hence, the amplification strategies are important for the SPR bioassay. In this thesis, a series of novel biosensing assay systems were developed for sensitive SPR detection based on aptamer-target recognition, bio-bar-code nanoprobe enhancement and enzyme assisted cycle amplification strategy. This paper research detailed content was mainly described as follows:1. A sensitive surface plasmon resonance method has been designed to selectively and sensitively detect lysozyme. In the present study, circular common target molecule displacement polymerization is developed to amplify the SPR signal for biomolecule assays, extending isothermal polymerization to an aptameric system without any medium. The isothermal circular polymerization-based bifunctional oligonucleotide probe facilitated the integration of multiple functional elements into a cooperative biosensing system:homogeneous assay format, isothermal signal amplification, aptameric target recognition, and hairpin DNA probe. In the presence of lysozyme, the target binding could induce the conformational change of the designed bifunctional probe that in turn triggered an isothermal circular polymerization reaction, contributing to the signal amplification. Utilizing this signaling scheme, the aptasensing system exhibited improved analytical properties. The detection limit of 1.68 pM could be achieved. The proposed biosensing scheme as a proof-of-concept was expected to promote the application of oligonucleotide probes in basic research and medical diagnosis.2. Detection of specific DNA sequences is important to molecular biology research and clinical diagnostics. To improve the sensitivity of surface plasmon resonance (SPR), a variety of signal amplification methods has been developed, including polymerase chain reaction (PCR) technology, molecular beacon, and AuNP enhancement. Herein, we described a new approach for nucleic acid detection by the combination of rolling circle amplification (RCA) reaction with SPR. Because of the binding of abundance repeated sequences of RCA products with gold nanoparticle probes, SPR signal was significantly amplified and the detection limit of 0.35 pM might be achieved. This RCA-based SPR might discriminate perfect matched target DNA from 1-base mismatched DNA with high selectivity. The high sensitivity and selectivity of RCA-based SPR makes it a potential tool for early diagnosis of gene-related disease and also offers a great promise for multiplexed assays with DNA microarrays.3. A sensitive and versatile surface plasmon resonance bioassay was proposed for the detection of DNA and Ramos cells by combining the target-triggered isothermal exponential amplification with the magnetic nanoparticle-based rolling circle amplification, producing remarkable amplification efficiency. The strategy for DNA assay included the following three steps:target-triggered isothermalexponential amplification reaction, magnetic nanoparticle-based RCA reaction, and AuNPs-enhanced SPR assay. This method exhibited a broad dynamic range, ultrahigh sensitivity and excellent specificity. The amplification strategy for cancer cells assay was proposed based on specific recognition of aptamers to cells. A detection limit of 10 cells was achieved. This assay represents a novel technique for both qualitative and quantitative detection of cancer cells in the blood sample, holding great promise for the broad applications in the field of medical research. |
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