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Experimental Study on the Detection of TORCH Infections Using Protein Microarray Technology

Author: YuZhangBin
Tutor: JiangLi
School: Southeast University
Course: Pediatrics
Keywords: protein microarray TORCH infections Biotin–Streptavidin System immuno–gold–silver staining
CLC: R714.7
Type: Master's thesis
Year: 2006
Downloads: 94
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Abstract


Objective: To search appropriate methods for modification and activation glass slides for protein immobilization,six different substrates were evaluated.By using gold nanoparticles and fluorescence as signal detection,the protein microarray for the detection of TORCH infections were researched.Methods: Six different methods for modification and activation glass slides were used,their contact angles were measured,atom force microscope were utilized to obtain surface profiles of the microarray on glass slides. We generated protein microarray by printing human IgM on modificative glass surfaces,the efficiencies、linear range、background of protein immobilizing on glass slides were compared;The antigens were printed on glass slide modified with the agarose films,the serum antibodies against the TORCH antigens were combined by using biotin labeled secondary antibodies. According to the affinity of Biotin-Streptavidin,the serum IgM antibodies of TORCH infections were detected;The antigens were printed on glass slide modified with the agarose films,the gold nanoparticles were introduced to the microarray by the specific binding of the gold-conjugated antibodies and then coupled with silver enhancement to enhance microarray signal, which could be easily imaged with a simple flatbed scanner or even the naked eye.Results: Their contact angles and surface profiles of the modificative glass surfaces,the efficiencies and the effects of immobilized proteins were discussed,the results indicated contact angles of glass slides modified with agarose films were bigger than the others, had much holes and thickness,demonstrated highest immobilization capacity、three dimension structure and the lowest signal background;Protein microarray based on Biotin-Streptavidin system and fluorescence as signal detection showed high detection sensitivity(0.5pg of IgM immobilized on glass slides);97.1% concordance was obtained between protein microarray and ELISAs;Protein microarray which are based on immuno–gold–silver staining method for the detection of TORCH infections demonstrated the low detection limit(12.5pg of IgM immobilized on slides);93.3% concordance was obtained between protein microarray and ELISAs in the classification of sera.Conclusion: The agarose films coated glass slide were appropriate substrates for protein immobilization of protein microarray;By using gold nanoparticles and fluorescence as signal detection,the protein microarray for the detection of TORCH infections were prepared,it is a high-throughput,low cost,rapid and simple approach;It was more practical and broad applications.

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CLC: > Medicine, health > Obstetrics and Gynaecology > Obstetrics > Perinatal medicine
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