Electrochemical Sensor Technology For Detection Of Pancreatic Cancer Specific Markers: K-ras Gene And Trypsin

Pancreatic cancer is a widespread alimentary canal cancer which has high fatality rate. When most of patients with pancreatic cancer seek medical advice, the pancreatic cancer is already terminal cancer which is unable to cure. The researches show that mutation rate of K-ras gene of patients with pancreatic cancer exceed90%. And the trypsin level of peripheral blood conspicuously increase in80%of patients with pancreatic cancer. Thus, mutant K-ras gene and abnormality of trypsin level in peripheral blood can act as the basis of making a definite diagnosis of early pancreatic carcinoma.In recent years, electrochemical biosensor technology is rapidly developing. The technology is playing an increasingly important role in genetic diagnosis, antibody detection and anti-cancer drug screening and so on. Combining the molecular pathology characteristics of pancreatic cancer, we designed the electrochemical sensor of diagnosing the early pancreatic cancer in the paper. The main contents are as follows:Chapter one constructing electrochemical DNA biosensor based on E.coli DNA ligase and AuNPs for the detection of mutant K-ras geneIn this section, construction the electrochemical DNA biosensor based on E.coli DNA ligase and AuNPs, RuHex acting as signal system, detected the mutant K-ras gene by the chronocoulometric. Using SEM and UV-VIS characterized the AuNPs and AuNPs probe, employing EIS characterized the preparation process. Under the optional experimenal condition, the linearity range of measuring mutant K-ras gene was from0.1pM to1000pM, and the LOD was10fM for mutant K-ras gene. The electrochemical DNA biosensor can accurately detect the mutant K-ras gene in the hybrid system containing the wild K-ras gene.Chapter two constructing electrochemical DNA biosensor based on DNA polymerase I for the precise detection of mutant K-ras geneIn this section, constructing electrochemical DNA biosensor based on DNA polymerase I, HRP immobilized on the modified electrode surface by the biotin-avidin syetem, precisely detected the mutant K-ras gene by the current-time curve technology in TMB solution. Using EIS characterized the preparation process. Under the optional experimental condition, the linearity range was0.001pM～100pM for mutant K-ras gene, and the LOD was0.5fM. The electrochemical DNA biosensor is superior to other method of detecting K-ras gene and accurately detected the K-ras gene in the hybrid system mixing with wild K-ras gene. That provided alternative method for detecting mutant K-ras gene in clinical.Chapter three constructing label-free electrochemical immunosensor based on PTh-AuNPs composite acting as signal system for the precise detection of trypsinIn this section, constructing label-free electrochemical immunosensor based on PTh-AuNPs composite acting as signal system, precisely detected trypsin by differential pulse voltammetry in the PBS with pH5.5. Using UV-VIS, TEM and XRD characterized the PTh-AuNPs, and employing AFM characterized preparation process of the immunosensor. Under the optional experimenal condition, the linearity range was0.02ng/ml-20ng/ml for trypsin, and the LOD was0.01ng/ml. The immunosensor can detect trypsin in serum of different pancreatic disease patient. The absolute value of relative standard deviation between the result and ELISA was less than9.0%. It was satisfactory. The label-free electrochemical immunosensor can measure the trypsin in serum in the clinical.