| Liver disease may have a serious effect on our health, and will have a greateffect on the social and economic development. Liver cancer is a malignant tumorwhich was one of the highest death rates in the world. Virus hepatitis, hepatic fibrosis,hepatocirrhosis and so on are the primary causes of liver cancer. So the diagnosis ofliver disease is an important part of the medical diagnosis. The human liver F proteinthat is not applied to clinical medicine is a new kind of biological mark which canreflect the extent of the liver damage directly. Since the 1980s, people began tostudy the relationship between the serum concentration of F protein and the extent ofthe liver damage, and obtained some clinical data. It will be used for a verypromising mark which can reflect the extent of the liver damage in high sensitivityand specificity by the great different concentration between the hepatocyte and serum,and also by the strict tissue specificity. Now medical study and application of liverspecific F protein is being paid more and more attention.This text is base on the study of the predecessor. Extracting the total mRNA ofthe human fresh liver tissue, after determining its purity and integrity, it was to beRT-PCR and PCR to amplify the purpose gene, then the purpose gene was connectedwith the pUCm-T carrier to obtain the cloning plasmid pUCm-T-F and putpUCm-T-F into the E.coli DH5a. The cloning part which was correct by sequencingwas connected with the expressing carrier pET-15b to obtain the expressing plasmidpET15b-F of F protein and put this expressing plasmid into expressing strainBL21(DE3)pLysS and used IPTG to induce its expressing. Purification was done byaffinity chromatograph (Ni2+-charged IDA His-bind column). The all stain proteinafter expressing was detected by SDS-PAGE and Western blot. And this protein wasused to immunize the rabbit to obtain the rabbit antihuman polyclonal antibody. Theindirect enzyme-linked immunosorbent assay was used to detect the titer of theantibody, and the Western blot was used to detect the specificity. The result of the cloning and expressing of human liver F protein showed thatthe size of the purpose gene was about 1200bp. This result was corresponded to theexpected value. The all stain protein after inducing and expressing by SDS-PAGEshowed that the size of purpose protein was 43kD. This result was corresponded tothe reports. The Western-blot showed that the strong immune reaction was occurredbetween purified protein and guinea pig antihuman extract F protein polyclonalantibody. This suggested that the purpose protein had the immunocompetence ofhuman F protein. The indirect ELISA to be used for the titer of the antibody, theresult suggested that the titer was 1: 6400, it could be used in immune reaction ofantigen and antibody. Western-blot suggested that the antibody prepared could bereacted with the extract human F protein, indicated that the rabbit antihumanpolyclonal antibody had the immunologic specificity, and it could be used for thecontinued study.The liver specific F protein was a sensitive effective and stable mark to reflectthe extent of the liver damage. And there was more important value in the earlydiagnosis and therapy about liver disease, drug screening and observation of curativeeffect, the condition of curing, judgement about extent of liver damage and forensicmedicine, the study on liver transplantation antirejection of immunotoleranee. It wasso difficult to obtain a great quantity of F protein by chemical extract, this preventthe other studies' progress. So, we obtained the gene cloning of human liver Fprotein by gene engineering, and induced the expression of human liver F protein,solve the problem in source. And this protein was used to immunize the rabbit toobtain the polyclonal antibody, and the Western blot was used to detect the specificity.That provided the foundation for the next study on trying to find out the optimumcondition of the immunoreagent, immunotolerance and immunological character.
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