Studies On The PAPS Diagnosis Reagent For T. Gondii's Antibody And The Surface Antigen 1 Expressed In E.coli

Toxoplasma gondii is an obligate intracellular protozoan parasite with an exceptional broad host range, also is the causative agent of toxoplamosis and responsible for serious zoonosis. In this study, the report refers to diagnosis methods and antigen for diagnosis, aimed to establish rapid and simple methods and to obtain better specific diagnosis antigen.1. Study on Polyaledehyde Polystyrene(PAPS) diagnosis reagent for T.gondii 's antibodyTo suit diagnosis on toxoplasis and quarantine animals on the ground floor, Latex Agglutination Test(LAT) treating PolysTyrene as vector was optimized. Polystyrene vector was connected with the functional group on the peaceful condition, which play an important role in vector attachment to protein, and the reaction caused to produce Polyaledehyde Polystyrene(PAPS). PAPS can adsorb protein in the model of not only physics but also chemistry, becoming more loosely than Polystyrene and can be immingled more easily, moreover, it's diameter gets wide. Storage phase showed that the stability of PAPS' functional group and it's combining protein didn't change in the nine month at four degree. Interesting, the sensitivity and stability of PAPS diagnosis reagent for T.gondii's antibody didn't change in the same state again. PAPS diagnosis reagent for T.gondii's antibody was contrasted with Indirect Heamagglutination Assay Kits on the market, the results demonstrated that the former was two dilution higher than the latter. It wasn't found that PAPS diagnosis reagent for T.gondii's antibody elicited the immune response with the positive serum of Schistosoma, Trypanosome, Trichina, Lung fluke, Fasciola hepatica, Bladder worm. PAPS diagnosis reagent and Indirect Heamagglutination Assay Kits reacted with the positive and negative serum simultaneously respectively, the results showed that the former and the latter was matched at detectable rate. The serums of two thousand and eight hundred dogs was investigated at Minhang District in Shanghai, the results indicated that the average positive rate is seventeen point three percent . To our knowledge, the methods can prolong the storage time of diagnosis reagent, and have better specificity, higher sensitivity, better repeatability and so on .2. Study on efficient expression of the truncated SAGl gene of Toxoplasma gondii in E.coli and identification of expression productThe surface antigen 1 protein (SAGl) is surface antigen of tachyzoite, which can stimulate body to generate antibody. At present, it is potently valuble for diagnose. To obtain the SAGl protein depending on molecular biology, the SAGl gene being truncated was expressed in E.coli BL21. Tachyzoite was collected from the abdomen of the mice by inoculating tachyzoite. A 993bp DNA fragment was amplificated by PCR directly from tachyzoite. The PCR product was cloned into pGEM-T easy plasmid by the way of T-A link, and the recombinant pGEM-T easy+SAGl plasmid was constructed after being transformed into E.coli DH5a. The truncated SAGl from recombinant plasmid after being cut by EcoRI and Hindlll constructed pGEX-2T+SAGl. After identification by restriction enzymes and sequence anylasis, the pGEX-2T+SAGl was transformed into E.coli BL21, and it was induced by IPTG. The best conformable IPTG concentration and induction time was...