A群脑膜炎奈瑟菌多糖IgG抗体检测试剂研制  

Development and application of an ELISA kit for detecting human anti-capsular polysaccharides IgG of serogroup A meningococcus

作　　者：肖丽君[1] 赵恩宏[2] 孔健[3] 郝志敏[2] 

机构地区：[1]承德医学院免疫微生物教研室,河北承德067000 [2]承德医学院附属医院 [3]北京绿竹生物技术有限责任公司

出　　处：《中国公共卫生》2008年第3期344-346,共3页Chinese Journal of Public Health

基　　金：国家科技部创新基金资助项目(03C26211100767)

摘　　要：目的研制检测A群脑膜炎奈瑟菌荚膜多糖IgG抗体的酶联免疫吸附试验（ELISA）检测试剂。方法以CTAB、酚、乙醇从A群脑膜炎奈瑟菌培养上清中提取荚膜多糖（MenA PS）,将多糖与卵清蛋白（EA）以还原氨基法偶联,并以Sepharose CL-4B纯化。优化反应条件,建立以偶联物MenA PS-EA为包被抗原检测A群脑膜炎奈瑟菌多糖IgG抗体的间接ELISA法,并与血清杀菌力试验（SBA）比较。结果高效液相色谱检测MenA PS-EA结合物纯度为81.50%。试剂具有良好重复性（变异系数CV=4.26%～5.30%）;37℃放置7d、4℃放置12个月,试剂的灵敏度不变,具有优良的稳定性;分别用该试剂和SBA法检测192份血清,试剂相对于血清杀菌力试验的灵敏度为96.91%,特异度为90.00%,一致性为95.83%,统计学分析表明,2种检测方法之间差异无统计学意义。结论以纯化MenA PS-EA作为包被抗原建立的A群脑膜炎奈瑟菌多糖特异性IgG抗体间接ELISA检测试剂具有可行性,为进一步研究及开发细菌多糖类抗体诊断试剂提供了可借鉴的方法。Objective Development and application of an ELISA kit for detecting human anti-capsular polysaccharides antibody of serogroup A meningococcus.Methods The capsular polysaccharide（MenA PS）was purified from the supernatant by CTAB,phenol,and ethanol precipitation.MenA PS was conjugated with egg albumin（EA）by reductive amination,and purified by Sepharose CL-4B.We optimized the reaction conditions,developed the indirect ELISA method for the detection of human anti-serogroup A meningococcus polysaccharide IgG which with the purified MenA PS-EA as coating antigen.The kit was compared with SBA.Results The purity of MenA PS-EA was 81.50% by high-performance liquid chromatography（HPLC）.The diagnostic kit was assessed by a series of assays,the sensitivity and specificity and reproducibility（CV of positive quanlity control sera：4.26%～5.30%）were good.It has an excellent stabilization when it was deposited at 37℃ for 7 days or 4℃ for 12 months.192 sera were tested by serum bactericidal assay（SBA）and ELISA to evaluate the sensitivity and specificity of the ELISA kit,when SBA was used as the gold criteria,the sensitivity,specificity of ELISA were 96.91%,90.00% respectively.The agreement between SBA and ELISA was 95.83%,no significant difference was found between the two methods by statistics.Conclusion The anti-MenA PS antibody ELISA kit was suitiable to measure human serum antibody level.Other anti-polysaccharid antibody ELISA kits may be developed according to this technic.

关 键 词：酶联免疫吸附试验(ELISA) 偶联物 抗A群脑膜炎奈瑟菌荚膜多糖IgG抗体 

分 类 号：R378.15[医药卫生—病原生物学]