Construction And Application Of Flexible Electrochemical Sensors For Depression Marker DCNP1 And Formononetin

Flexible electrochemical sensors have the advantages of simple sample pretreatment,fast response,light weight,great portability,high sensitivity,and strong adaptability to the environment,which further expand the application of sensors in the analysis of biomarkers and active components of traditional Chinese medicine.The global lifetime prevalence of depression is about 16%,and it has become the fourth most common disorder in the world.However,the current diagnosis and treatment evaluation of depression mainly rely on subjective scales,lacking the assistance of objective methods.Dendritic cell nuclear protein-1（DCNP1）is a potential marker of depression,of which quantitative analysis may be helpful in the early diagnosis of depression.Formononetin,an active ingredient widely present in Chinese medicine such as Radix Astragali,has various pharmacological effects.It is of great significance to achieve its sensitive,rapid,and highly selective detection in complex systems for the further study of formononetin.To achieve the selective and sensitive determination of DCNP1 and formononetin,the electrochemical sensing technique was employed in this article,using a flexible electrode（ITO-PET）as the base electrode with nitrogen-doped graphene（NG）and its composite（NG/TiO₂）prepared with titanium dioxide nanoparticles（TiO₂）to amplify the response signal.A label-free flexible immunosensor based on NG/TiO₂and a molecularly imprinted flexible sensor based on NG were successfully constructed and applied to determine the depression marker DCNP1 in rat brain homogenate samples and formononetin in traditional Chinese medicine extracts and plasma samples,respectively.The details are as follows:1.Label-free flexible immunosensor based on NG/TiO₂ for the determination of depression marker DCNP1DCNP1 is a newly discovered potential marker of depression,and its sensitive determination is of great significance for the early diagnosis and treatment evaluation of depression.In this work,a label-free flexible immunosensor for selective and sensitive detection of DCNP1 was successfully constructed.At first,a highly conductive nanomaterial NG/TiO₂ was prepared using a one-pot method,and it was used as an electrode modification material in the construction of immunosensors for the first time.NG/TiO₂ was functionalized with 1-pyrene-butyric acid N-hydroxysuccinimide ester（PSE）,and then the DCNP1 antibody could be better immobilized on the surface of the modified ITO-PET.Then bovine serum albumin（BSA）was used as a blocking agent to block non-specific active sites,and the immunosensor（BSA/anti-DCNP1-PSE/NG/TiO₂/ITO-PET）can be obtained.The fabricated NG/TiO₂ and the construction process of the immunosensor were characterized by scanning electron microscopy,infrared spectroscopy,cyclic voltammetry,and electrochemical impedance method.Different concentrations of DCNP1 were measured by differential pulse voltammetry under optimized experimental conditions.The constructed sensor has a wide linear range of1～10~4pg/ml,a low detection limit of 0.24pg/ml（S/N=3）,with good reproducibility,selectivity,and stability.And it has been successfully used for the determination of DCNP1 in rat brain homogenate samples,providing a possible method for the rapid and accurate determination of DCNP1,as well as the early diagnosis and treatment evaluation of depression.2.Molecularly imprinted flexible sensor based on nitrogen-doped graphene for highly sensitive and selective determination of formononetinFormononetin,an isoflavone compound widely present in Chinese medicine such as Radix Astragali and Radix Puerariae,has various pharmacological activities,including anti-inflammatory,anti-cancer,and hypoglycemic effects.It is of great significance to achieve its sensitive,rapid,and highly selective detection in complex systems for the in-depth study of formononetin.In this work,a molecularly imprinted flexible sensor was constructed.At first,NG with better electrochemical performance was prepared from graphene oxide,and the ITO-PET was modified.Then,using formononetin as the template and o-phenylenediamine as the functional monomer,electropolymerization was carried out to form a molecularly imprinted polymer（MIP）on the electrode surface.And the molecularly imprinted flexible electrode（MIP/NG/ITO-PET）was obtained after elution.Scanning electron microscopy,Raman spectroscopy,infrared spectroscopy,cyclic voltammetry,and electrochemical impedance method were used to characterize the electrode morphology and sensor construction process.Effects of modification amount,polymerization conditions,elution conditions on sensor performance were studied.Under the optimized experimental conditions,the molecularly imprinted flexible sensor can selectively detect formononetin in the linear range of 3～120μM,with a detection limit as low as 1.14μM（S/N=3）,good reproducibility,stability,and anti-interference ability.And it has been successfully applied in the analysis of formononetin in the methanol extract of Radix Astragali and mouse plasma with good accuracy and precision.In summary,a label-free flexible immunosensor（BSA/anti-DCNP1-PSE/NG/TiO₂/ITO-PET）for selective and sensitive detection of DCNP1 and a molecularly imprinted flexible sensor（MIP/NG/ITO-PET）for selective analysis of formononetin were successfully constructed in this article.Both of them have a fairly wide linear range and low detection limit,with good reproducibility,selectivity,and acceptable stability.And both have been successfully used for the analysis of target analytes in real samples,providing possible new methods for selective and sensitive detection of DCNP1 and formononetin in complex systems.