Comparison Of Surface Modification For Protein Microarray And Application Research

Purpose:Surface modification of protein microarray carrier is one of the key factors affecting quality of microarray, protein microarray carriers not only requires high performance fixed target protein, and requires that the target protein with strong activity can capture the target protein. Until now, for surface modification of protein microarray carrier is no uniform point of view. Papers on the basis of summing up the predecessor, compared several common methods for surface modification of microarray carrier, the purpose is to compare the different methods of modification of glass on the merits of the protein efficiency. Also, explore a better cosmetic methods used to prepare antigen microarray, and preparation of antigen microarray for detection of four tumor markers of liver, verify that the antigen microarray on feasibility of combined multiple tumor markers antibody detection, preliminary study on its application.Methods:Using molecular self-assembly technology, glass slides were modified with (3-aminopropyl) trimethoxysilane, glutaraldehyde, poly-L-lysine to prepare protein microarrays. Evaluate and choose the optimal surface modification methods for the preparation of protein microarray. The immobilizing proteins were labeled by Cy3, attachment rate was evaluated. Then, the mouse IgG were printed on the modified glass slides, the target proteins were labeled by Cy3, responsive activity of protein on microarrays were evaluated. For AFP, CEA, CA19-9 and Fer were printed on the modified glass slides, using the principle of indirect ELISA to detect the samples including AFP, CEA, CA19-9, and Fer antibodies, the purposes is to detect four andibodies of liver tumor marker at the same time. Protein microarray incubation is completed, washing; tested the reaction fluorescence intensity by ScanArray Gx biochip scanner, preparation methods for surface modification of protein chip for better discussion and adoption of protein microarray for detection of multiple tumor markers.Results:At the protein attachment rate, the attachment rate of target protein is between 0.8 and 1.0 after the glutaraldehyde modification of protein microarray carrier, most of the target protein attachment rate of poly-1-lysine and amino-silane is less than 0.5; At the responsive activity, when the concentration of Cy3 labeled antibody is less than 5μg/mL, after the glutaraldehyde modification of glass is higher than poly-1-lysine and amino-silane modified glass slides, after the glutaral-dehyde modification preparation of protein microarray carrier with strong ability to capture free complementary proteins.Using protein microarray detect antibodies of four tumor markers of liver, can be achieved simultaneously with different types of antibody in the sample for qualitative analysis, required less sample.Conclusion:This research has established protein microarray surface modification method in the laboratory, has established the better surface modification methods of protein microarray carriers, the aldehyde modification have higher background, have a larger detection limit, the reactive activity of the immobilized proteins has also demonstrated more superior. Using the glutaraldehyde modification of preparation of antigen microarray slide, can be used for combined detection of multiple tumor markers antibody. A case study of detection of four tumor markers antibody of liver, preliminary validation of the feasibility; provides data reference to the development of unmarked antibody-antigen complexes detection system in the late.