Biosensing Design Based On Nucleic Acid Probes And Nanoenzymes And Its Application In Cancer Biomarkers Detection

The high fatality rate of cancer is a serious threat to human life and health,and the current techniques of cancer diagnosis have certain limitations.The appearance and development of cancer biomarkers has brought new opportunities for early cancer diagnosis.The rapid development of biosensing technology provides a new research platform for the creation of efficient,ultra-sensitive,highly selective,and highly accurate cancer biomarker detection methods,and has important theoretical and practical significance for protecting human life and health and reducing medical costs.The flexible structure and strong specific recognition ability of nucleic acid probes and nanozymes combining the unique structure and function of nanomaterials and biological enzymes offer a brand-new method and idea for constructing novel and diverse biosensor technology.This paper is based on nucleic acid probes and nanomaterials to exploit a high-sensing biosensing detection platform and take advantage of fluorescence,electrochemical detection techniques to explore its application in the detection of cancer biomarkers,which has important research significance on the early diagnosis of cancer.First,this paper presented a ratiometric fluorescent nucleic acid probe to detect the telomerase activity of cancer biomarkers in urine of bladder cancer patients by combining the characteristics of nucleic acid probes and fluorescent dyes.Herein,two non-interfering fluorescent dyes were selected to improve the reproducibility and robustness.One was red emissive aggregation-caused quenching(ACQ)dye,Cy5,as a control.The other was blue emissive aggregation-induced emission(AIE)dye,Silole-R,as reporter of telomerase activity.In the presence of telomerase,the telomerase primer chains extended so that the blue emission was enhanced by the added negatively charged sites for Silole-R to bind and aggregate,while the red emission was almost unchanged as stable internal reference.With the addition of incremental amounts of telomerase,the ratiometric emission intensity ratios(I478/I665)of this bioprobe gradually increased.Furthermore,the distinguishing of telomerase extracts from 20 bladder cancer bloody and 10 normal urine specimens confirmed the practicality of this bioprobe.The ratiometric fluorescent nucleic acid probe utilizes a stable internal reference to achieve high reproducibility,high sensitivity,and high selectivity detection of the telomerase of the cancer marker,and is expected to be applied in clinical diagnosis.Second,this paper developed a flexible microelectrode modified by dual nanozymes based on graphene quantum dot(GQDs)assemblies and Au Pd alloy nanoparticles(Au Pd-ANPs)using a new type of material that has enzyme characteristics and nanomaterial properties,namely nanozymes and the unique advantages of electrochemical method,and explored its practical application in electrochemical sensing system for sensitive detection of cancer biomarker hydrogen peroxide(H2O2)in human breast cancer cells and tissue.For the preparation of dual nanoenzyme modified microelectrode,ionic liquid was used as the electrolyte for the effective electrodeposition of GQDs on carbon fiber substrate to form a close-packed assembly under a very negative potential,then the highly dense and ultra-fine Au Pd-ANPs were uniformly decorated on GQDs assembly by electrodeposition.In virtue of the unique structural merits and synergistic contribution of dual nanoenzyme in enhancing the electrocatalytic activity to H2O2,the resultant nanohybrid microelectrode exhibited good sensing performances for electrochemical detection of H2O2,including a high sensitivity of 371 μA cm-2 m M-1,a wide linear range from 1.0 μM to 18.44 m M,a low detection limit of 500 n M(a signal-to-noise ratio of 3:1),as well as good selectivity and biocompatibility,which could be used for real-time tracking H2O2 released from different types of human breast cells and in situ sensitive detection of H2O2 in clinical breast cancer tissue.This will be of great application value for the in vitro and in vivo detection of implantable and wearable electrochemical sensing devices.