关键词: 弓形虫, 免疫印迹, 试剂盒, 敏感性, 特异性

Abstract: Objective To develop a diagnostic kit for the detection of anti-Toxoplasma antibodies in human sera by immunoblot technique. Methods The cytoplasm proteins of Toxoplasma gondii were extracted from collected ascites in Kunming strain mice infected by tachyzoites of Toxoplasma gondii(RH strain). Toxoplasma gondii soluble antigens were separated by SDS-PAGE electrophoresis and transferred to pyroxylin membrane. Using nonpoisonous tetramethylbenzidine （TMB） as horseradish peroxidase substrate for immunoblot， the optimal experimental conditions were selected through comparison of antigen preparation， reagents for blocking or washing， dilution concentration， reaction time， and frequencies of reacting band. Sensitivity， specificity， Youden index and stability were evaluated as the standard for the kit. Results In 30 sera with anti-Toxoplasma IgM antibodies and 28 with IgG antibodies， the sensitivity for IgM and IgG antibody detection was 90.0% （27/30） and 85.7% （24/28） respectively， the specificity was all 100% in examining 40 healthy control sera， and the Youden index was 0.9 and 0.86 respectively. The kit was stable at 4℃ for 6 months. Conclusion The immunoblot kit shows an easy operation， fast reaction and reliable result， and may be practical.

Key words: Toxoplasma gondii, Immunoblot, Kit, Sensitivity, Specificity