The Study Of Integration Protein Chip In The Detection Of Toxoplasma Gondii Infections

Protein chips also named protein microarrays ,they are small flat surfaces that allow highly parallel analysis of thousands of molecular parameters in a single experiment. Recently, protein chips attracted more and more attention because of its potentially important advantages in throughput. Furthermore, the concept of protein arrays offers the potential advantage of being scalable, flexible, automatable and easy to perform. The advantages of the technology make the use of protein arrays extend into many areas. In this thesis, A new method of protein arrays was developed for detecting Toxoplasma gondii infections, and spotting concentration, immobilizing time ,target labeling and affinity reaction condition were investigated. The major contributions include the following aspects:1. We modified carriers(glass slides)with agarose films to allow antigens to be immobilized on them during the preparation of protein chips, established the Toxoplasma gondii antigen concentration on the spotting,the immobilizing time,the incubating time which specifically interact between antigens and antibodies.We also labeled two kinds of the second antibodies with fluorescein Cy3.2. On the basis of the antigen-antibody interaction priciples, we detected the immobilization efficiency of antibodies IgM, IgG, and created the Log[IgM,IgG]-flourescence intensity curves which had a good linear correlation between the signal intensity and the the antibodies concentration, we ascertained the most proper labor concentration of antibodies IgM,IgG immobilizing on agarose films as masculine comparision.3. Protein microarrays for detection of Toxo. gondii antibodies in human sera were fabricated by printing antigens on agarose film coated glass slide. The antibodies in the criterion serum and clinic samples against the Toxo. gondii antigen were detected by using fluorescence labeled secondary antibodies. The Toxo . gondii assay performed equivalently to traditional ELISA Method, but in smaller sample and reagent volumes, shorter reaction time and higher sensitivity.So we can draw a conclusion that the protein microarray have the advantages of high throughput, micromation, continuum , speediness, accuracy and so on, which will be used in clinic in the future.