ZnO Micro-/Nano Array Substrate Consturction And Their Performance In Tumor Marker Detection

With the entensive in-depth application of nanomaterials in the field of biomedical detection,the development of low-cost,high-sensitivity and good biocompatibility nanomaterials has become an urgent need in the medical industry.The traditional biological detection method has the disadvantages of low sensitivity,high cost and long time-consuming.In addition,the reagents used in traditional biological detection（such as fluorescent molecules,etc.）usually have a short life time and a weak signal output.In the detection of biomarkers,precious metal materials（gold,silver,etc.）are widely used,such as fluorescence detection,Raman detection,and immune indications.The large-scale promotion of precious metals is largely limited by production costs and fabrication processes,and it is unstable in some applications（such as Raman detection）.Therefore,for noble metal materials,the shortcomings of high cost,low stability and poor repeatability are still needed to be solved in practical applications.In contrast,ZnO stands out because of advantages such as simple preparation process,low cost,and stable signal output（such as fluorescence enhancement applications）.ZnO has been applied in many fields of biological detection,such as fluorescent enhancement substrates for biomarker detection,Raman enhanced substrates for small molecule detection and micro/nano structure-based substrates for capturing circulating tumor cells（CTCs）,etc..However,due to the defects of ZnO itself（strong non-specific adsorption,emission easy to be quenched etc.）,ZnO needs further functionally modified before being applied in practice.This work trys to solve the above problems by functionalization of ZnO.Microfluidic chemical synthesis method is used to prepare ZnO substrates.The surface characteristics of ZnO are functionalized with organic or inorganic modification strategies to enhance the detection performance of the substrate,or by adjusting its structure growth to obtained the three-dimensional（3D）micro-nano structure for CTC capture,etc.The details are as follows:1.Based on the fluorescence enhancement characteristics of ZnO,an ultra-sensitive CEA detection system was constructed in the glass capillary channel using a hydrophilic modification strategy.First,the growth mechanism of ZnO nanorods in microchannels was studied,and it was found that ZnO nanorod arrays with high aspect ratio have higher fluorescence enhancement ability.Subsequently,after hydrophilic modification by layer-by-layer self-assembly,the ZnO nanorod array showed an effective anti-nonspecific adsorption ability.Finally,after grafting the specific antibody,the minimum concentration of 0.1 pg/mL carcinoembryonic protein（CEA）can be detected within 1 h.The microfluidic device has the potential to be used as a point-of-care diagnostic（POC）tool and applied in daily health monitoring.2.On the basis of ZnO fluorescence enhancement,in-situ coating of the fluorescence enhancement material can realize the double amplification effect of the substrate on the fluorescent molecules.First,the ZnO nanorod array was successfully constructed in the microchannel by the batch reaction method,and then the array with the best fluorescence enhancement performance was optimized.After demonstrating that ZIF-8 has a fluorescence enhancing effect on fluorescent molecules,the ZnO nanorod array combined with CEA and fluorescent molecules was coated in situ with ZIF-8.Under the double amplification effect,CEA can be detected in the range of0.01–100 pg/mL within 1 h.The microfluidic device shows more sensitive detection performance for CEA.3.On the basis of the ZnO electron transfer characteristics,the ZnO nanorod array is combined with the two-dimensional Ti₃C₂T_x nanosheets to achieve highly sensitive detection of the analytes through the electron transfer effect.First,a microfluidic chemical synthesis method was used to prepare the inclined ZnO nanorod array,and the two-dimensional Ti₃C₂T_x solution was deposited on the surface of the ZnO array by the drop-casting method to obtain a ZnO/Ti₃C₂T_x substrate.After optimization to obtain the substrate with the best SERS performance,the detection performance of rhodamine 6G（R6G）was comparable to that of the noble metal substrate,and the detection limit of miRNA reached 10^-7 M.Density functional theory（DFT）simulations and calculations have found that the interface charge transfer between ZnO and Ti₃C₂T_x and between Ti₃C₂T_x and rhodamine 6G（R6G）molecules greatly contributes to the improvement of detection sensitivity.The preparation method is simple and has excellent detection performance.Highly sensitive detection performance was obtained for small organic molecules without the use of noble metals,which provides a new idea for the development of non-noble metal SERS substrates.4.Taking advantage of the easy controllability of the ZnO structure,the efficient capture and controllable release of CTC are realized by constructing a three-dimensional（3D）hierarchical structure array in the microchannel with polymer functional modification.First,a 3D hierarchical Zn（OH）F/ZnO nanoforest array was constructed on the inner wall of the glass capillary,and then the temperature-sensitive polymer-poly（N-isopropylacrylamide）（PNIPAAm）was successfully modified using the"grafting from"strategy.After further conjugate modification of CTC-specific antibody anti-EpCAM,the capture efficiency of CTC in whole blood samples was as high as 81%.In addition,the introduction of PNIPAAm reduces the non-specific capture of cells by the substrate and increases the cyto-compatibility of the substrate.This method provides the idea of simple preparation of 3D hierarchical structure and the strategy of interface organic functionalization of ZnO.