The Study On Menthods Of Separation And Detection Of Glycoprotein Based On Novel Boronate Affinity-material

Glycoproteins,one of the most important products of protein post-translational modification,are involved in many complex biological processes.Studies have shown that abnormal expression levels of glycoproteins are closely related to the occurrence of many diseases,so specific analysis of glycoproteins is of great significance for the early diagnosis and real-time monitoring of certain diseases.However,the low abundance of glycoproteins and the interference of other matrices in complex biological systems have brought great challenges to the research of glycoproteins.Therefore,it is important to efficiently separate and enrich glycoproteins before analysis.This study mainly focuses on the separation and enrichment of glycoproteins.Two nanomaterials based on boronic acid affinity were prepared for efficient separation of glycoproteins and a molecularly imprinted resonance light sensor was constructed to detect glycoproteins with high sensitivity on this basis.The details are as follows:1.A boronic acid affinity-magnetic hydroxyapatite was designed and synthesized for separation and enrichment of glycoproteins.First,polyethyleneimine was coated on the surface of magnetic hydroxyapatite by electrostatic interaction,and then phenylboronic acid was covalently grafted by amide reaction.The prepared nanomaterials have high binding ability to glycoproteins,and the adsorption amounts to ovalbumin(OVA),transferrin(TFR)and horseradish peroxidase(HRP)are 586.43 mg/g,457.57 mg/g and 527.57 mg/g,respectively.2.A novel Wulff-type boronic acid modified magnetic metal-organic frame imprinting polymer is prepared for specific recognition of ovalbumin under physiological conditions.First,the magnetic metal-organic frame is used as a substrate,and then gold nanoparticles(Au NPs)were grown on the surface of the substrate by electrostatic interactions.Second,covalently graft Wulff-type boronic acid molecular group onto the surface of nanoparticles,and the target ovalbumin was then immobilized on the surface of the nanoparticles by boronic acid affinity.Finally,the imprinted layer is formed by self-polymerization of tetraethyl orthosilicate,and the template is then eluted under acidic conditions to obtain an imprinted polymer.The experimental results show that the imprinted polymer has excellent binding capacity(337.8 mg/g)and high selectivity(IF=4.52)for ovalbumin under physiological conditions.The adsorption equilibrium is reached when the adsorption time is 20 minutes.3.A magnetic metal-organic frame molecularly imprinted resonance light sensor based on boronic acid affinity was proposed to detect glycoproteins.After the sensor is combined with the target glycoprotein,it can cause the change of resonance light intensity.The sensor has a wide linear range(0.05-1.8 n M)and low detection limit(13 pM)for glycoproteins.