| Drug development and food safety are issues of vital relevance to human health and development.Traditional drug development is time-consuming,low success and cost-intensive.A scanty few candidates will eventually pass FDA approval.In order to improve the success rate and reduce the cost of drug development,the accuracy of pharmacologic research,especially pharmacodynamic analysis,in the pre-clinical research stage has a significant impact on the efficiency and success rate of drug development.In the aspect of Marine food safety,the increasingly serious environmental pollution leads to more aquatic products containing Marine biological toxins.The accumulation of Marine biological toxins through the food chain seriously threatens human health and public safety.Therefore,it is of great significance to study the high accuracy drug-screening sensor and the high sensitivity Marine biological toxin detection sensor.This paper was based on biosensor technology,aiming at the demand of high-precision and high-sensitivity sensors for drug screening and toxin detection in the field of life and health,and carried out research on 3D cell-based biosensor for drug screening and molecular biosensor for toxin detection.Based on 3D cell-based biosensor,this paper developed 3D Electric Cell-substrate Impedance Sensing(ECIS),designed two kinds of 3D ECIS chips,established corresponding detection methods and models,and applied them to the screening of anti-tumor drugs.At the same time,based on the molecular sensing technology and using highly specific immune molecules and aptamers as sensitive components,two kinds of highly sensitive Marine biological toxin detection sensors were established and applied to the analysis of actual shellfish samples.The main research contents and innovative work of this paper include:1.3D Electric Cell-substrate Impedance Sensing(3D ECIS)modle was estabilished,and 3D ECIS chip was designed to realize the dynamic monitoring of the growth status of 3D cells.In this paper,a real-time,non-invasive 3D ECIS was developed for monitoring 3D cell growth status and drug sensitivity.According to the characteristics of 3D cell culture in non-conductive matrigle,the equivalent circuit model of 3D cell-based biosensor was established and verified by software simulation.The total impedance value of the cell and gel mixture was analyzed with the impedance detection technology.A new method for dynamic monitoring of the growth state of 3D cells was proposed to monitor the growth process of 3D liver cancer cells in real time and the changes of cell activity after the action of anti-tumor drugs.The experimental results show that 3D ECIS can be effectively used to evaluate the growth status and efficacy of 3D cells,and it is a promising platform for anti-cancer drug screening.2.A 3D micro-groove impedance sensor(3D MGIS)was designed and fabricated based on micro-nano technology to realize real-time monitoring of 3D cell growth status and high-throughput drug screening.In this paper,a 3D microcavity impedance sensor(3D MGIS)with higher stability and throughput was designed and fabricated using MEMS technology for dynamic monitoring of 3D cell growth.According to the changes of the 3D MGIS chip,the working parameters were optimized,the equivalent circuit model was adjusted,and the simulation verification was performed again.The effect of cisplatin,gemcitabine and pemetrix were evaluated by 3D lung cancer model and MGIS chip.The experim ental results showed that compared with 2D ECIS,3D ECIS can effectively distinguish the efficacy differences of lung cancer drugs and improve the accuracy of drug screening.3.A method for detection of shellfish toxin based on portable flow cytometry combined with magnetic bead immunosensor was proposed,it has the advantages of high sensitivity,high precision and fast detection speed.The biotinylated Okadaic acid(OA)modified on the surface of magnetic beads and in the solution to be tested,OA competitively reacted with OA antibody.Subsequently,the second antibody labeled with algin amplified the signal,and the field fluorescence intensity of the immune magnetic beads was analyzed by portable flow cytometry.The detection range of the sensor for OA is 0.5?20 ?g/L,and the detection limit is 0.05 ?g/L.With the introduction of magnetic beads,the complexity of experimental operation is reduced,the detection time is effectively shortened,and the anti-interference ability of the sensor is significantly improved.Compared with traditional mouse biological method and liquid chromatography,this method has the characteristics of fast detection speed,high accuracy and good portability.The test results of the actual samples show that the sensor has the potential to be used in the field rapid detection of OA toxin.4.A method for detection of shellfish toxin based on aptamer biosensor was proposed,it has the advantages of high stability,good specificity and low cost.In order to meet the demand of high stability and low cost for field detection of shellfish toxin,an aptamer-based biosensor was constructed.In this paper,the gold nanoparticles were modified on the surface of the silanized forked finger electrode,and then the OA aptamer was fixed on the forked finger electrode by electrostatic adsorption.OA adaptor will block the catalytic active sites of gold nanoparticles.After the combination of OA and OA aptamer in the solution to be tested,the activation sites of the gold nanoparticles were released and began to self-catalyzed growth,causing the conductivity of the forked finger electrode to rise.The conductance of the cross finger electrode was analyzed by linear sweep voltammetry,and then OA was analyzed and measured.The OA detection range of the sensor is 5?80 ?g/L,and the detection limit is 1 ?g/L.As a recognition element,the aptamer has lower cost,better stability and stronger affinity compared with the antibody.Combining with the electrochemical workstation with higher versatility,this method is easier to be extended to first-line use. |
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