Preparation And Primary Application Of Immunity Colloidal Gold Test Strip For Toxoplasmosis

Toxoplasma gondii(T.gondii)disease is a zoonosis and more than 200 kinds of animals can be infected with the disease,most of them show recessive infection,but infected pig can result in large numbers of death.Toxoplasmosis also can cause great harm for pregnant women and immunocompromised persons;Toxoplasmosis is serious to threaten human’s health and causes considerable economic losses in livestock.Currently,the clinical diagnosis of toxoplasmosis mainly rely on serological diagnosis,including indirect hemagglutination test,indirect fluorescent antibody test,dye test and enzyme-linked immunosorbent assay.But these immunoassay techniques have distinct drawbacks which make it unsuitable for use in the field,These methods require special equipment and the procedures are time-consuming.Colloidal gold technology is a new rapid diagnostic techniques.For its easy operation,rapid determination,no relevant professional and technical personnel,no expensive equipment needed,simple clinical detection colloidal gold technology has been developing rapidly in recent years.Now clinical diagnosis of toxoplasmosis mainly detect the serum antibodies.But the detection of serum antibodies can not distinguish between current infection or previous infection.In this study,a new diagnostic method to detect circulating antigen of Toxoplasma gondii was established by using immunological methods,labeling and chromatography technique.Tow new monoclonal antibodies which were made by our laboratory were used in establishing the new method.Firstly the monoclonal antibodies of hybridoma cell lines D5 and B6 against Toxoplasma tachyzoites antigen and P30 antigen and Toxoplasma tachyzoites antigen were prepared and purified.Simultaneously,the colloidal gold particle was prepared,and then the affinity purified D5 monoclone antibodies was labeled with the colloidal gold particle.The R6 monoclonal antibodies and the goat anti-mouse IgG antibody were coated on the surface of nitrocellulose filter membrane(NC)as the test line(T line)and control line(C line)respectively.By a series of optimizing the expenriment conditions,the immune colloidal gold test strip was developed.The sensitivity test showed the strip can detected in the lowest dilution of 1:1000 Toxoplasma circulating antigen.And it has no cross reaction with the antigens of Trpanosonma evansi’VSG,Fasciola gigantica’ES,Metagoninmus Yokogawai,Anisakis.64 circulating antigen in serum of mice,which is from established animal model of mice infecting with Toxoplasma gondii,were detected by the test strip and in the meantime double antibody sandwich ELISA method was established for comparison.The results showed the total coincidence rate with the tow methods was 93.8%;80 porcine serum collecting from Guangxi farm also detected by these two methods and the total coincidence rate was 98.8%,the test result is highly consistent.The above results showed the detecting efficacy of the new method has highly consistent to the double antibody sandwich ELISA method.The experiment results proved that this diagnostic test strip is a sensitive,specific,rapid and accurate method for detecting Toxoplasma gondii’s circulating antigen.The establishment of immune colloidal gold test strip has provided a new diagnostic method for current infection of Toxoplasma gondii and it has very important sense for more efficacious prevent and treatment of toxoplasmosis.