Multi-functional Cardiomyocyte-based Biosensor Systems And Their Applications In Pharmaceutical Analysis And Biotoxin Detection

Drugs and food,essentials of everyday life,are closely connected to the survival and development of mankind.Drugs,which should be the representative of safety,efficacy and quality controllability,still may cause adverse drug reactions occasionally during normal use resulting in drug withdrawals all over the world.For discovery and precaution of drug induced side effect,rigorous drug safety evaluation,especially the cardiac safety examination of drugs should be operated during the preclinical studies.Organism produced biotoxins hiding in our food occur to be another serious threat to our health in daily life.Due to the accelerated toxin production induced by the heavy environmental pollution,disease-causing biotoxins can be found throughout the human food chain.Therefore,studies on safety assessment of drugs and food are of great significance.Up to date,cell-based bioassays act as alternative and practical in vitro biological assays for drug screening and biotoxin detection.In this thesis,a multifunctional cell-based impedance biosensor system and a cardiomyocyte-based hybrid-biosensor system are proposed for application of drug screening in preclinical studies and marine biotoxins detection combining the technique of cell-based biosensor with cardiomyocytes.The main contents and innovations of this thesis are given as follows:1.A new approach for simultaneous detection of the growth and mechanical beating of cardiomyocytes with a newly developed cell-based impedance biosensorCell-based impedance biosensor reflects the cellular morphology changes including cell attachment,migration and death on the surface by alternating current impedance technique.In this thesis,a multifunctional cell-based impedance biosensor is proposed for detection of not only the growth but the mechanical beating status of cardiomyocytes.The cell-based impedance biosensor is optimized by the cell seeding density experiments and verified by the cellular growth and beating experiments of cardiomyocytes.2.Effects and cardiotoxicity assessments of cardiovascular drugs using a new detecting instrument for simultaneous detection of cardiomyocyte growth and mechanical beatingToxicology researches about drugs,especially the cardiotoxicity researches,in the preclinical studies are of great importance in the drug discovery procedure.In this thesis,the multifunctional cardiomyocyte-based impedance biosensor provide a multi-dimensional analysis for effects and cardiotoxicity assessment of cardiovascular drugs by use of growth and mechanical beating monitoring of cardiomyocytes.A voltage-sensitive calcium channel activator,bay K8644,an L-type calcium channel blocker,verapamil,and a slow delayed rectifier potassium channel blocker,chromanol 293B are used for functional verification of cardiac effects and safety assessment.3.A new approach for detection of the viability and electric excibility of cardiomyocytes utilizing a newly developed cell-based hybrid-biosensor integrated with interdigitated electrodes and planner electrodesThe integration and combination of different kinds of biosensors is one of the developing trends in field of cell-based biosensor.In the thesis,the interdigitated electrodes are integrated with planner electrodes for construction of the hybrid-biosensor.Cardiomyocyte is coupled with the newly developed hybrid-biosensor,providing a comprehensive portrait of the cardiomyocyte model via short-term electrophysiological recording and long-term viability monitoring in a same culture.The cardiomyocyte-based hybrid-biosensor is optimized by the cell seeding density experiments and verified by the growth and electrophysiology monitoring experiments of cardiomyocytes.4.Chemotherapy drugs test and marine biotoxins detection using a novel detecting instrument for cardiomyocyte viability and electric excibility monitoring.The toxicity of drug and food is a serious issue in field of health care and it is also the common application area of cell-based biosensor.In this thesis,the cardiomyocyte-based hybrid-biosensor is construct with cardiomyocytes and the integration sensor of interdigitated electrodes and planner electrodes.It provides a comprehensive analysis of the chemotherapy drugs including taxol and vinblastine by growth and electrophysiology monitoring of cardiomyocytes.Furthermore,a typical toxin of diarrhetic shellfish poisoning,okadaic acid and a typical toxin of paralytic shellfish poisoning,saxitoxin are detected respectively according to the specificity of cytotoxicity and electrophysiological toxicity.