Agarose gel-based protein microarray immunosensor preparation method based on function of maleimide and application

Abstract

The invention relates to an agarose gel-based protein microarray immunosensor preparation method based on the function of maleimide and application of an agarose gel-based protein microarray immunosensor and belongs to the field of electrochemical sensing and immunoassay. According to the method and application of the invention, an organic synthesis method is adopted; agarose gel (AG) is activatedby using 3-Maleimidobenzoic acid N-hydroxysuccinimideester (MBS); and stable thioether bonds are formed by means of specific recognition between the C-terminal cysteine sulfydryl of a recombinant protein A and unsaturated carbonyls in the MBS, so that a protein-based microarray is constructed. With the method adopted, the stability of the secondary and tertiary structures of the protein A can beeffectively maintained; an N terminal capable of recognizing the Fc domain of an antibody is fully exposed, so that the activity of an immobilized antibody can be enhanced; an electrochemiluminescenceimmune sensing strategy is adopted, so that the ultra-sensitive detection of a disease marker can be realized. The sensor has the advantages of simplicity in construction, low cost consumption, highsensitivity, wide detection range and great clinical application potential.

Description

technical field [0001] The invention belongs to the field of electrochemical sensing and immune analysis. Background technique [0002] The sensitivity of electrochemical immunosensors depends on the characteristics of the substrate material cured on the surface of the sensor, the type of antibody, the number of immune molecules and its remaining activity. Although the use of nanomaterials as the sensing matrix has achieved good results, many materials such as cadmium-based The inevitable biological toxicity of the compound and the intricate electron transfer path greatly affect the repeatability of the sensor; [0003] Staphylococcus aureus protein A is a surface protein evenly distributed on the cell surface of Staphylococcus aureus, it has a high affinity with different globulins, especially the five highly homologous IgG binding domains at the N-terminal can specifically It recognizes the antibody Fc fragment in human serum, and at the same time, the wide range of non-b...

AI Technical Summary

Problems solved by technology

[0002] The sensitivity of electrochemical immunosensors depends on the characteristics of the substrate material cured on the surface of the sensor, the type of antibody, the number of immune molecules and its remaining activity. Although the use of nanomaterials as the sensing matrix has achieved good results, many materials such as cadmium-based The inevitable biological toxicity of the compound and the intricate electron transfer path greatly affect the repeatability of the sensor;