Photocatalytic Electrosensor For Label-free Detection Of Breast Cancer 1 Gene And Lysozyme

With the development of industry,environmental pollution problem has become more and more serious.With their improvement of living standards,people increasingly concerned their health.Particularly,people pay more attention to these diseases caused by environmental pollution.Therefore,developing a rapid,sensitive and reliable disease-related detection method is very urgently.In recent years,more and more people focus on the determination of DNA,protein and small molecules.There are numerous detection methods including fluorescence,chemiluminescence,photocatalytic visualization,etc.Electrochemical biosensor has been used widely due to the advantages of fast response,high sensitivity,simple design,low cost.In this work,we found that the dye molecules interacted with DNA could photocatalytically generate singlet oxygen(1O2)and thus cleaved the DNA upon light emitting diode(LED)irradiation,leading to an obvious change of [Fe(CN)6]3-/4-redox current.Therefore,two photocatalyticly electrochemical sensor for label-free detection of breast cancer 1(BRCA1)gene and lysozyme(LZM)were fabricated.The main contents of this thesis are as follows.(1)Photocatalytic electrosensor for label-free and ultrasensitive detection of BRCA1 gene in serum was developed.The capture DNA(DNA-c)modified with thiol group was immobilized on the gold electrode through thiol-Au interaction.Upon recognition of target DNA(DNA-t)by DNA-c,the ethidium bromide(EB)molecules that were embedded in the hybridized double strand DNA(dsDNA)could photo-catalytically generate 1O2,leading to an efficient cleave of the dsDNA.As a result,the electrochemical signal of [Fe(CN)6]3-/4-increased remarkably because of less blocking of electrode and lower charge repulsion.Such a simple strategy provided an ultrasensitive detection of BRCA1 gene.The factors of electrochemical photocatalytic system were systematically investigated,including the choice of electrolyte system,the optimization of dyes,the choice of LED lamps,the concentration and the incubation time of EB,the LED illumination time.Under the optimum experiment conditions,the beoad linear range for BRCA1 was achieved from 10 aM-100 nM withdetection limit of 1.7 aM.The photocatalytic electrosensor was the most sensitive for detection of BRCA1 gene by far in comparison of those without an amplification procedure.Also the sensor can discriminate mismatched DNA from perfectly matched target DNA with high selectivity.Two serum samples spiked with 100 pM and 100 aM BRCA1 gene were analyzed.The meanrecoveries ranged from 87.0% to 95.3%,indicating that the proposed sensor can be employed as a reliable method for BRCA1 gene detection in real samples.Therefore,free lable,simplicity,high sensitivity,high selectivity and specificity provided by this sensor will make it a promising tool for early diagnosis in gene-related diseases.(2)Photocatalytic electrosensor ultrasensitive detection of LZM in urine based on graphene oxide(GO)and Phloxine B(PB)was developed.First,when the LZM aptamer were hybridized with DNA-c on the gold electrode surface.Upon recognition of LZM by capture LZM aptamer,dsDNA unwinded into ssDNA.GO adsorbed the DNA-c modified on electrode.Then PB was combined with GO through ?-? stacking.The PB molecules could photo-catalytically generate 1O2,leading to an efficient cleave of the DNA.As a result,the interfacial electron-transfer resistance of [Fe(CN)6]3-/4-decreased.Such a simple strategy provided an sensitive determination of LZM.The experimental conditions were systematically optimized,including the optimization of dyes,the concentration and the incubation time of GO,the concentration and the incubation time of PB and the LED illumination time.The hexagonal cells structure of the GO was used to combine the PB,and GO with spccific surface area can amplifed the detection signal.Coupled with photocatalytic and electrochemical detection methods,sensitivity was improve greatly.Under the optimum experiment conditions,the concentration of LZM was linear from 1 pM-100 nM and the detection limit was 140 fM.Thrombin,bovine serum,human serum,adenosine triphosphate,glucose,myohemoglobin,carcino-embryonic antigen and VEGF were chosen to investigate the selectivity of the present biosensor.Finally,the method is applied to the detection of urine spiked samples.The recoveries were from 91% to 101%.Therefore,free label,simplicity,high sensitivity provided by this sensor will make it a promising tool for disease detection.