Study On Cortisol Aptamer Sensor Based On Surface Plasmon Resonance Enhancement By Gold Nanoparticles

Cortisol is the main active substance of glucocorticoid in human body,which plays an important role in regulating various physiological processes such as blood pressure,glucose level and carbohydrate metabolism.Abnormal concentration of cortisol can cause clinical symptoms such as obesity,osteoporosis,blood pressure and blood sugar disorders.Therefore,it is of great significance to develop a highly sensitive and fast real-time response cortisol sensor for human health monitoring.The conventional cortisol detection technique uses the method of cortisol antibody recognition,which has the disadvantages of high immunogenicity and instability.Compared with antibody recognition,aptamers(single stranded DNA or RNA nucleotides)with high affinity and high selectivity can effectively overcome the above shortcomings of antibody detection,providing a simpler processing method and a lower cost detection method for cortisol detection.Gold nanoparticles(AuNPs)have the characteristics of large specific surface area,low biotoxicity and easy to be modified.The local surface plasmon resonance(LSPR)effect of AuNPs is increasingly used in biosensors.In this thesis,cortisol aptamer with strong recognition ability was selected as the ligand and an AuNPs-Apt conjugate cortisol probe was prepared to achieve quantitative detection of cortisol.The coupling enhancement effect of AuNPs on SPR sensor was used to improve the sensitivity of SPR sensor,and then a novel SPR cortisol sensor was constructed,while the preparation conditions and sensing characteristics of the sensor were studied indetailed.In addition,combining with the advantages of quick response,small size,low cost and strong anti-interference ability of optical fiber sensor,a novel optical fiber cortisol sensor was prepared by combining optical fiber sensor technology with AuNPs,and the preparation process of the sensor was optimized,finally realizing the real-time detection of cortisol.The main contents are as follows:(1)AuNPs with different particle sizes were prepared by the seed method,and AuNPs-Apt coupling was prepared by the salt aging method.The structure and morphology of AuNPs and conjugated compounds were characterized by UV-Vis,FTIR and SEM,respectively.The cytotoxicity of AuNPs-Apt conjugate was tested by the CCK-8 method.(2)Cortisol aptamers can specifically recognize and bind to cortisol,and the AuNPs-Apt coupling probe sensor was constructed by modifying the aptamers with activated sulfhydryl group(-SH)onto the surface of AuNPs through Au-S bond.The preparation conditions of the sensor were optimized by adjusting the concentration of NaCl during the preparation process,and the performance of the sensor in the detection of cortisol solution was studied.The sensor has a detection range of 1-1000 n M and a response time of 80 minutes.(3)A SPR cortisol sensor was constructed by combining the LSPR effect of AuNPs with SPR enhancement.AuNPs were fixed on the surface of 50 nm thick gold film with 1,6-hexadimercaptan as the crosslinking agent.The surface of AuNPs was modified with cortisol aptamer.The effect of preparation conditions on the performance of the sensor was systematically studied,including the particle size of AuNPs and the incubation time of AuNPs on the gold film.The results show that the sensitivity of the sensor improved by 45% after the AuNPs modification.After biofunctionalizing the sensor with cortisol aptamer,the standard curve,detection range and selectivity of the sensor for cortisol solution detection were studied.(4)The AuNPs-Fiber LSPR sensor was prepared using polyallylamine hydrochloride(PAH)as the crosslinking agent and AuNPs electrostatic self-assembly monolayer was formed on the side of the exposed fiber core.The effects of AuNPs particle size,incubation time and core diameter on the performance of optical fiber sensor were investigated.The results show that the RI sensitivity of the optical fiber LSPR sensors is 609.09 nm/RIU under the optimal preparation conditions.In this thesis,a novel fiber-optic cortisol sensor was constructed by using the surface biofunctionalization of AuNPs with cortisol aptamers.The standard curve,detection range and selectivity of the sensor for cortisol solution detection were studied,and this sensor realized the fast and real-time detection of cortisol was realized.