The Construction And Applification Of An Antifouling Graphene Transistor Biosensor For The Detection Of Disease Biomarker EpCAM

The mortality rate of tumor diseases has remained high,mainly due to the metastasis of malignant tumors.If early diagnosis and treatment can be achieved,the 5-years survival rate of patients would be greatly improved to 90%,even 100%for some cancers.Among the many tumor detection methods,liquid biopsy has attracted a lot of attention because of its non-invasive sampling,which can directly detect circulating tumor cells（CTCs）in the patient’s blood to assess the condition.However,there are problems and challenges due to that the biomarkers of liquid biopsy are low concentrations,need for enrichment,and high cost in the detection process.Epithelial cell adhesion molecule（EpCAM）is a glycosylated transmembrane protein,which is highly expressed on the surface of CTCs corresponding to liver cancer,esophageal cancer,gastric cancer,etc.,and participates in the adhesion and migration of CTCs to promote tumor metastasis,so EpCAM is recognized to be as a biomarker for CTCs in liquid biopsy.In summary,the development of a highly sensitive,highly specific,simple and efficient EpCAM detection method is of great significance for biological research and early clinical diagnosis.The field effect transistor is a kind of electronic sensor device that can amplify the signal,is highly sensitive,miniaturizable and easy to integrate,which havee attracted the attention of researchers in many fields such as chemistry,food,environment,and medical treatment in recent years because of its high performance.Solution-gate field effect transistors can provide low voltage（-1 V-1 V）analysis and detection,which may avoid the electrolysis of molecules and maintain bioactive dielectric layer solution.This is very conducive to the analysis and detection of bioactive molecules and provide guarantee for the smooth diagnosis of various disease markers,which has attracted the favor of the majority of researchers.The advantages of high mobility and bipolarity of graphene materials make graphene field-effect transistors attract much attentions in the field of biological detection.However,there are many disadvantages in the preparation of graphene,such as the high price of CVD method,some toxic raw materials in chemical reduction method,long reaction time,and the mechanical peeling method without guarantee of efficiency and quality,which cannot meet the needs of industrial preparation and integration of transistor sensing.Therefore,the method study of graphene preparation with efficient,fast and green on the surface of channel in transistor is very important for the industrialization of graphene transistor biosensing.In this work,an electrodeposition method was studied in detail on the surface of transistor channel,and solution-gate graphene transistor biosensor was constructed for the highly specificity and sensitive detection of cancer biomarker EpCAM,which provided a powerful in vitro diagnostic method for early diagnosis of cancer.（1）Electrodeposition reduction of graphene oxide（ERGO）to prepare solution-gate graphene transistors.Firstly,poly-L-lysine was used to improve the adhesion of graphene in the glass interface in order to improve the stability of the transistor.Then,three different electrodeposition routes were studied including one-step 1,one-step 2 and two-step.It was found that two-step electrodeposition was better and favorable for performance for the transistor.The electrodeposition conditions were also investigated in detail such as electrolyte concentration,p H value,scanning speed,scan cycles and GO amount etc.Moreover,the carrier mobility of the solution-gate ERGO transistor was estimated to beμ_p=20.3 cm²V^-1s^-1,μ_n=6.3 cm²V^-1s^-1.The studies provided a green,non-toxic,fast and efficient method to fabricate graphene field effect transistors,which was a great advantage for mass-product sensor with low-cost.（2）Construction and application of ERGO transistor biosensor with the carbon nanocomposite film and its quantitative detection of EpCAM.In real samples of biomarkers,there are usually some interfering substances,which will attach to biosensor and even affect the detection results of targets.Graphene,in spite of its high isoelectric property of electron mobility,however,has a strong adsorption effect on many substances,which is a deficiency that must be overcome in the analytical detection of complex environmental disease biomarkers.Physical isolation methods can passivate channel materials effectively sequestering various macromolecules.Here,a functionalized carbon nanocomposite film was designed to prevent the nonspecific adsorption of graphene in transistor biosensor and improve the sensing specificity.Bovine serum albumin（BSA）,glutaraldehyde（GA）and carboxylated carbon nanotube were firstly used to form a nano three-dimensional structure,which was drop coated on the graphene channel to form a protective film to prevent solution environment nonspecific adsorption during the detection process.The results indicated that this carbon nanocomposite membrane can effectively prevent non-specific adsorption and had antifouling properties.Based on the shift of the electroneutral point（Dirac point）of the characteristic curve of the transistors transfer,EpCAM was detected quantitatively.The results showed an excellent linearity in the range of 50 pg/m L-50 ng/m L withΔV_CNP=11.2836-9.8399lgc（R²=0.9959）and the detection limit of 0.24 pg/m L.These studies have shown that the transistor biosensor can provide highly specific and sensitive detection of EpCAM in complex solution environments.