Research Of Outer Surface Precise Functionalized Nanochannels And Their Applications In Biosensing

It is of great significance to accurately detect these bio-markers for understanding biological processes.The solid-state nanochannels based sensing systems has been demonstrated with advantages in detection of bio-markers,due to the rapid,label-free,and high-throughput screening with high sensitivity and specificity.However,traditional solid-state nanochannels research focus on the probes on their inner wall（IW）,ignoring probes on their outer surface（OS）.As a result,the sensitivity of traditional solid-state nanochannels for bio-markers detection still need to be further improved.Recently,research on solid-state nanochannels with probes on their outer surface has been demonstrated their ability of independent regulating-ion-transport.This finding not only contributes to in situ detect bio-markers with large size but also provides promising opportunities for detection with ultra-high sensitivity and clear mechanism.In this work,a series of solid-state nanochannels sensing systems with different functionalization of the OS were constructed by accurately partitioning IW and OS of nanochannels.The effects of wettability of the IW and OS,distribution of positive and negative charges on the OS,and probes density on the OS of nanochannels on the ion current signal were systematically studied,which provided a new idea for the development of efficient solid-state nanochannels detection system.The main research contents are as follows:（1）The wettability of the IW and OS of solid-state nanochannels on its detection performance was studied.Nanochannels with hydrophilic DNA probes at the inner wall(DNA@IW_Hydrophilic)and hydrophobic coating at the outer surface(None@OS_Hydrophobic)were constructed by modifying fuctional elements with precise partition.The synergistic effect between None@OS_Hydrophobic and DNA@IW_Hydrophilic,and its structural-activity relationship with detection sensitivity were systematically studied.Compared with the traditional solid-state nanochannels with hydrophilic probes distributed on both the IW and OS,the nanochannels with DNA@IW_Hydrophilic+None@OS_Hydrophobic significantly decrease the limit of detection（LOD）by 10⁵-fold,the LOD of Hg²⁺as low as 1 n M was realized.Through molecular dynamics simulation,the mechanism of improving the sensitivity of None@OS_Hydrophobic was further explored.Meanwhile,nanomolar detection of Hg²⁺in vivo fish muscle was achieved.This work provides important insights into the development of high sensitivity and high reliability solid-state nanochannel detection sensors in complex samples.（2）The charge distribution on the OS of nanochannels on the accurate discrimination of multiple proteins was studied.Three solid-state nanochannels were first prepared,（1）symmetrical nanochannels with the same charges on IW and OS;（2）asymmetrical nanochannels with the same charge on IW and OS;（3）asymmetrical nanochannels with opposite charges on IW and OS.Three kinds of solid-state nanochannels discrimination analysis systems were constructed by modifying the DNA probes on the OS of the nanochannels for discrimination of multiple proteins.The effect of nanochannels with different charges on IW and OS on ion current signals was systematically explored.They achieved 78.3%,95.0%and 100.0%accurate discrimination of bovine serum albumin（BSA）,casein（Cas）,cytochrome C（CC）,human serum albumin（HSA）,horseradish peroxidase（HRP）,and lysozyme（Lyso）,respectively.Meanwile,the reliability of the solid-state nanochannels discrimination analysis system was verified for the discrimination of 6 proteins in artificial serum.This work provides theoretical guidance for the design of nanochannels discrimination analysis platform with space charge asymmetry.（3）The probes density on the OS of nanochannels on the accurate discrimination of multiple proteins was studied.Firstly,graphene oxide（GO）was filtered to form a nanochannels matrix.Then,based on the properties of temperature regulating the intermolecular/intramolecular interactions of bioconjugation at different interfaces,DNA1 and DNA2 probes were driven to bind to regions with different wettability on the OS of GO nanochannels at-20℃,room temperature,70℃,respectively.A series of nanochannels array with different density of DNA probes on the OS were constructed.The temperature-controlled bonding force between the probe and the OS of the nanochannels on the graft density of the probes was investigated,and the relationship between graft density of probes and ion current signal before and after target binding was systematically studied.The three nanochannel arrays were used to discriminate 7 proteins,including BSA,Cas,CC,HSA,HRP,Lyso and fibrinogen（Fib）with an accuracy of90.0%,94.3%and 100.0%,respectively.Among them,the solid-state nanochannel array prepared at-20℃successfully realized the discrimination of 7 proteins in artificial serum.This work provides new insights for designing temperature-controlled nanochannels arrays with different probe densities on the OS.