An Electrochemical Biosensor Based On Graphite-like Phase Carbon Nitride For Ultra-sensitive Detection Of MiRNA-155

According to the World Health Organization,breast cancer has become the most common cancer worldwide and a serious threat to human life and health.Micro RNA（miRNA）,a class of endogenous non-coding single-stranded small RNA molecules,with miRNA-155 as one of the most widely studied miRNAs,its high expression during disease development makes it a novel biomarker for breast cancer.Objective To achieve sensitive,specific and rapid miRNA-155 detection is a challenge due to the complexity of human blood fractions and low miRNA-155 content.In this study,a novel miRNA-155 electrochemical sensor was constructed to meet the requirements of trace detection of the target,by taking advantage of the emerging electrochemical sensing technology and nanomaterials for sensitivity.Initially,to explore the possibility of miRNA-155 application to assist in clinical diagnosis of breast cancer.Method This approach designs an electrochemical biosensor based on graphite-like phase carbon nitride nanocomposites and hairpin-type capture probes for ultrasensitive detection of miRNA-155.The nanocomposites were synthesized by using carbon nitride nanosheets（g-C₃N₄）,poly-D-glucosamine（PDG）and gold nanoparticles（Au NPs）,modified on the glassy carbon electrode（GCE）surface.And the experimentally processed hairpin capture probes（HP）were prepared as bioreceptors for specific recognition of miRNA-155.In addition,the nanomaterials and electrode surfaces were characterized by electron microscopy,UV-absorption spectroscopy,linear cyclic voltammetry（CV）and electrochemical impedance（EIS）.Experimental conditions including nanocomposite ratios,nanomaterial modification volume,capture probe concentration,capture probe incubation time,and serum dilution times have also been optimized.Finally,the designed miRNA-155 biosensor will be used for the detection of miRNA-155 standard solution,working solution and actual serum samples by applying differential pulse voltammetry（DPV）.Regarding the performance of this miRNA-155electrochemical biosensor with terms of accuracy,reproducibility,specificity and stability,it was then validated by a series of methodological experiments designed.Results Electron microscopy,UV-absorption spectroscopy and electrochemical analysis were used to characterize the materials and electrode surfaces,demonstrating the successful preparation of the nanocomposites and their structural and functional integrity.The current signal was significantly amplified by its modification on the electrode surface.After the optimized experiments,DPV measurements of miRNA-155 standard curve and working curve were performed.An excellent linear relationship was found between the current signal difference（ΔI）and log of miRNA-155 concentration,the standard curve equation was calculated as y（ΔI,μA）=6.141x（lg C[miRNA-155],mol/L）+115.186,with the limit of detection（LOD）as low as 0.05 f M.Performance evaluation experiments such as spiked recovery,reproducibility,specificity and stability also showed promising results.In addition,measurements of actual serum samples from healthy people and breast cancer patients with different states found different current differences in different categories of actual samples,indicating different levels of miRNA-155.Conclusion The miRNA-155 electrochemical biosensor designed in this study achieved ultra-sensitive detection of the target in the linear range of 10^-16～10^-12mol L^-1and was initially applied to actual serum samples for detection.Thus,the preliminary exploration of this study in real samples shows its potential to assist in clinical diagnosis and screening.