中华微生物学和免疫学杂志
中華微生物學和免疫學雜誌
중화미생물학화면역학잡지
CHINESE JOURNAL OF MICROBIOLOGY AND IMMUNOLOGY
2011年
11期
1031-1034
,共4页
韩金乐%贾继宗%李川%杨亮%何德磊%李娟%由鹏飞%叶祥忠%李益民
韓金樂%賈繼宗%李川%楊亮%何德磊%李娟%由鵬飛%葉祥忠%李益民
한금악%가계종%리천%양량%하덕뢰%리연%유붕비%협상충%리익민
肠道病毒71%双抗体夹心ELISA%特异比活
腸道病毒71%雙抗體夾心ELISA%特異比活
장도병독71%쌍항체협심ELISA%특이비활
Enterovirus 71%Double antibody sandwich ELISA%Specific activity
目的 建立肠道病毒71型(EV71)抗原的ELISA检测方法.用于EV71的抗原定量、TCID50试验及EV71免疫原性与免疫保护性的关系分析.方法 以高亲和力的EV71中和单抗K8G2和Y8H2-HRP构建检测EV71抗原的双抗体夹心ELISA方法.比较本方法与显微镜观察法检测的EV71 TCID50.分析EV71抗原的特异比活与免疫血清中和抗体水平的关系.结果 本试剂定量检测抗原的线性范围为0.125 ～4.0 U/ml,R2 =0.9911,试剂不与EV71之外的受试物反应,试剂的回收率为0.89 ～ 1.16,变异系数小于15％,37℃9 d的热回收率大于85％.本法与显微镜观察法检测的EV71 TCID50的相关系数r=0.990.21株EV71抗原的特异比活与免疫血清中和抗体水平的相关系数r=0.930.结论 构建了EV71抗原ELISA检测试剂,可用于EV71的抗原含量检测、TCID50分析,为特异比活与免疫血清中和性关系研究提供了一些有价值的信息.
目的 建立腸道病毒71型(EV71)抗原的ELISA檢測方法.用于EV71的抗原定量、TCID50試驗及EV71免疫原性與免疫保護性的關繫分析.方法 以高親和力的EV71中和單抗K8G2和Y8H2-HRP構建檢測EV71抗原的雙抗體夾心ELISA方法.比較本方法與顯微鏡觀察法檢測的EV71 TCID50.分析EV71抗原的特異比活與免疫血清中和抗體水平的關繫.結果 本試劑定量檢測抗原的線性範圍為0.125 ～4.0 U/ml,R2 =0.9911,試劑不與EV71之外的受試物反應,試劑的迴收率為0.89 ～ 1.16,變異繫數小于15％,37℃9 d的熱迴收率大于85％.本法與顯微鏡觀察法檢測的EV71 TCID50的相關繫數r=0.990.21株EV71抗原的特異比活與免疫血清中和抗體水平的相關繫數r=0.930.結論 構建瞭EV71抗原ELISA檢測試劑,可用于EV71的抗原含量檢測、TCID50分析,為特異比活與免疫血清中和性關繫研究提供瞭一些有價值的信息.
목적 건립장도병독71형(EV71)항원적ELISA검측방법.용우EV71적항원정량、TCID50시험급EV71면역원성여면역보호성적관계분석.방법 이고친화력적EV71중화단항K8G2화Y8H2-HRP구건검측EV71항원적쌍항체협심ELISA방법.비교본방법여현미경관찰법검측적EV71 TCID50.분석EV71항원적특이비활여면역혈청중화항체수평적관계.결과 본시제정량검측항원적선성범위위0.125 ～4.0 U/ml,R2 =0.9911,시제불여EV71지외적수시물반응,시제적회수솔위0.89 ～ 1.16,변이계수소우15％,37℃9 d적열회수솔대우85％.본법여현미경관찰법검측적EV71 TCID50적상관계수r=0.990.21주EV71항원적특이비활여면역혈청중화항체수평적상관계수r=0.930.결론 구건료EV71항원ELISA검측시제,가용우EV71적항원함량검측、TCID50분석,위특이비활여면역혈청중화성관계연구제공료일사유개치적신식.
Objective To develop an ELISA method for quantitative determination of enterovirus 71 (EV71) antigen.The method can be applied to detect EV71 antigen contents and analyze the correlation between immunogenicity and immunoprotection.It also can be used for tissue culture infective dose( TCID50 ) assay.Methods A double antibody sandwich ELISA method was developed for quantitative determination of EV71 antigen on the basis of the high-affinity neutralizing monoclonal antibodies ( K8G2 and Y8H2-HRP).This method was compared with microscopic observation for the detection of EV71 TCID50.The correlation was analyzed between the specific activity of EV71 antigen and the EV71 neutralizing antibody titer in immune serum.Results The linear range of this method was 0.125-4.0 U/ml and the R2 value was 0.9911.The reagent did not react with other antigens except EV71 antigen.The recovery ratio of this method was 0.89-1.16.The coefficient of variation was less than 15％.The heat recovery rate was above 85％ when the reagent was in 37℃ for 9 days.There was a good correlation in TCID50 of EV71 between this method and microscopic observation,r=0.990.The specific activity of EV71 antigen had positive correlation with the neutralization titer of immune serum in 21 EV71 strains,r=0.930.Conclusion The quantitative ELISA method for EV71 antigen was developed,which could be used to detect EV71 antigen contents and analyze TCID50.The specific activity of EV71 antigen detected by the method could be used to evaluate the immunoprotection of the vaccine potency test.
