Screening And Functional Study Of HIV P24Aptamers&I-PCR Test P24HPV Serological Diagnoses Study

HIV infection is currently widespread in the world, becoming the twenty-first century’s most significant public health events. Current HIV resulting human AIDS (Acquired Immunodeficiency Syndrome, AIDS) remains medical problem, the existing treatments are too expensive, resistance to the toxic side effects. With a tremendous amount of financial resources and researchers to carry out vaccine studies, to date, no clinically effective products are available. AIDS is obviously not only a medical problem, but also social issues and economic issues. In the present case, the early detection, early diagnosis and early treatment of HIV is clearly the most important part. The prediction of the spread of infection and the evaluation of the drug treatment effectiveness require specificity and high sensitivity diagnosis. Currently, the detection of HIV infection by several ways, the p24antigen and antibody detection is the most commonly used clinical methods for HIV infection.p24antigen is a viral major structural proteins of HIV, a molecular weight of24kilodaltons (KD). p24antigen of HIV capsid of the main components, each of the complete virus particles contain about2000p24antigen. Early in HIV-infected individual, p24antigen has been expressed in the human body, and can be detected in the blood. Then the anti-p24antibodies appear in the blood, because the antigen-antibody neutralization, p24difficult to detect anti-out principle. Current methods for the detection of the p24ELISA method using ELISA method, can be detected in plasma as low as lOpg/ml, but this is bound to the generation of false negative. Therefore, it is necessary to further improve the detection sensitivity. The Immuno-PCR technology is undoubtedly by far the most sensitive detection methods, In theory, the Immuno-PCR method can greatly improve the detection sensitivity, which can shorten the window period of HIV caused AIDS, and more is mainly used to detect effects of the drug, and for the detection of blood products can greatly improve the sensitivity and reduce the testing costs. The currently used HIV nucleic acid detection methods, in particular the HIV viral load test, are very expensive, therefore Immuno-PCR method using the indirect determination of the amount of HIV viral load may be a partial substitution, in order to provide effective prevention and treatment of AIDS.SELEX (Systematic Evolution of Ligand by Exponential Enrichment) is a new type of in vitro screening technologies invented20years ago. The combined library technology and screening technology are used to find out a section of nucleic acid molecules from its library:ligand-aptamers; aptamers are similar to antibodies binding with specific target substance. SELEX aptamer technology has also been applied to biomedicine and other aspects. SELEX has also been considerable progressed, aptamers related to various viruses have been screened out, providing a new way for the prevention and treatment of AIDS-related disease.The first part of this thesis using SELEX technology for HIV-1p24antigen aptamer screening; study the screened aptamer affinity and specificity; and applying the aptamers to HIV-positive blood samples and healthy controls by Immuno-PCR assay to determine the p24antigen.Human papillomavirus (HPV) is double-stranded DNA virus, is the first virus to be identified as the causative agent of human cancer, especially the female cervical cancer. Currently more than200HPV types have been found, according to its risk of induce human tumors are divided into high-risk and low-risk. In China mainland, type HPV16and HPV18is the most important high-risk viral type. The majority of cervical cancer and other tumors with these two types are closely related.For virus-related tumors, serological diagnosis of nasopharyngeal carcinoma is firstly established by the academician Zeng Yi through the EB-related antibodies, creating a highly effective method for early screening of carcinoma, achieving outstanding contributions for prevention and control of nasopharyngeal cancer. Logically, for the DNA tumor virus HPV, we could also be able to find the appropriate serological diagnostic marker for the population screening and early diagnosis of HPV-associated malignancies. Therefore, the second part of this thesis is about serological detection of HPV infection. The target are mainland prevail high-risk HPV16, HPV18type. E6E7gene of the two high-risk HPV16and HPV18were constructed to prokaryotic and eukaryotic expression vector respectively. E6E7antigen was stably expressed. Purified E6E7antigen was used to detected antibodies from blood samples of clinical cancer.