中华地方病学杂志
中華地方病學雜誌
중화지방병학잡지
Chinese Journal of Endemiology
2014年
5期
498-503
,共6页
杜春红%王鹏%张建中%高子厚%尹家祥%张正飞%李寿芹%谭红丽%宋志忠
杜春紅%王鵬%張建中%高子厚%尹傢祥%張正飛%李壽芹%譚紅麗%宋誌忠
두춘홍%왕붕%장건중%고자후%윤가상%장정비%리수근%담홍려%송지충
鼠疫%重组F1抗原%胶体金免疫层析试纸条%应用评价
鼠疫%重組F1抗原%膠體金免疫層析試紙條%應用評價
서역%중조F1항원%효체금면역층석시지조%응용평개
Plague%Recombinant F1 antigen%Gold immuneochromatography assay%Field evaluation
目的 表达和纯化重组鼠疫菌F1抗原(rF1),构建胶体金免疫层析试纸条(GICA),检测和评价该试纸条的现场应用效果.方法 以镍离子金属螯合(Ni-NTA)亲和层析柱和脱盐柱对rF1进行纯化并构建GICA,应用GICA、双抗原夹心酶联免疫吸附试验(ELISA)和间接血球凝集试验(IHA)3种方法对1份鼠疫菌免疫兔血清和1 685份现场血清样本(包括94份人、939份犬、34份猫、609份鼠和9份旱獭)进行F1抗体检测.结果 rF1得到高效表达并实现较好纯化.GICA、ELISA、IHA 3种方法检测1份鼠疫菌免疫兔血清,结果均为阳性,最低检测滴度ELISA法高于GICA法(1∶10×211比1∶10×29),GICA法高于IHA法(1∶10×29比1∶10×28).对1 685份现场血清样本进行检测,GICA法阳性89份(5.28％),IHA法阳性53份(3.15％),ELISA法阳性101份(5.99％);GICA法与IHA法的总符合率为97.3％,与ELISA法的总符合率为97.9％.配对资料x2检验结果显示,GICA法阳性检出率高于IHA法(x2=26.63,P＜0.05),但与ELISA法比较,差异无统计学意义(x2=3.36,P＞0.05).结论 rF1表达和纯化成功,用其构建的GICA特异性强,敏感性高,与IHA、ELISA法结合应用,能有效提高鼠疫检测的速度和质量.
目的 錶達和純化重組鼠疫菌F1抗原(rF1),構建膠體金免疫層析試紙條(GICA),檢測和評價該試紙條的現場應用效果.方法 以鎳離子金屬螯閤(Ni-NTA)親和層析柱和脫鹽柱對rF1進行純化併構建GICA,應用GICA、雙抗原夾心酶聯免疫吸附試驗(ELISA)和間接血毬凝集試驗(IHA)3種方法對1份鼠疫菌免疫兔血清和1 685份現場血清樣本(包括94份人、939份犬、34份貓、609份鼠和9份旱獺)進行F1抗體檢測.結果 rF1得到高效錶達併實現較好純化.GICA、ELISA、IHA 3種方法檢測1份鼠疫菌免疫兔血清,結果均為暘性,最低檢測滴度ELISA法高于GICA法(1∶10×211比1∶10×29),GICA法高于IHA法(1∶10×29比1∶10×28).對1 685份現場血清樣本進行檢測,GICA法暘性89份(5.28％),IHA法暘性53份(3.15％),ELISA法暘性101份(5.99％);GICA法與IHA法的總符閤率為97.3％,與ELISA法的總符閤率為97.9％.配對資料x2檢驗結果顯示,GICA法暘性檢齣率高于IHA法(x2=26.63,P＜0.05),但與ELISA法比較,差異無統計學意義(x2=3.36,P＞0.05).結論 rF1錶達和純化成功,用其構建的GICA特異性彊,敏感性高,與IHA、ELISA法結閤應用,能有效提高鼠疫檢測的速度和質量.
목적 표체화순화중조서역균F1항원(rF1),구건효체금면역층석시지조(GICA),검측화평개해시지조적현장응용효과.방법 이얼리자금속오합(Ni-NTA)친화층석주화탈염주대rF1진행순화병구건GICA,응용GICA、쌍항원협심매련면역흡부시험(ELISA)화간접혈구응집시험(IHA)3충방법대1빈서역균면역토혈청화1 685빈현장혈청양본(포괄94빈인、939빈견、34빈묘、609빈서화9빈한달)진행F1항체검측.결과 rF1득도고효표체병실현교호순화.GICA、ELISA、IHA 3충방법검측1빈서역균면역토혈청,결과균위양성,최저검측적도ELISA법고우GICA법(1∶10×211비1∶10×29),GICA법고우IHA법(1∶10×29비1∶10×28).대1 685빈현장혈청양본진행검측,GICA법양성89빈(5.28％),IHA법양성53빈(3.15％),ELISA법양성101빈(5.99％);GICA법여IHA법적총부합솔위97.3％,여ELISA법적총부합솔위97.9％.배대자료x2검험결과현시,GICA법양성검출솔고우IHA법(x2=26.63,P＜0.05),단여ELISA법비교,차이무통계학의의(x2=3.36,P＞0.05).결론 rF1표체화순화성공,용기구건적GICA특이성강,민감성고,여IHA、ELISA법결합응용,능유효제고서역검측적속도화질량.
Objective To establish a gold immuneochromatography assay(GICA) using expressed and purified recombinant F1 antigen(rF1) of Yersinia pestis and to evaluate its field application effect.Methods rF1 was purified by Ni-NTA affinity chromatography column and desalting column and the GICA was established using purified rF1.Plague F1 antibody of one immune rabbit serum and 1 685 field serum samples(including 94 human,939 dog,34 cat,609 rat and 9 Marmot serums) were detected using GICA,double antigen sandwich enzymelinked immunosorbent assay (ELISA) and indirect hemagglutination assay (IHA).Results rF1 was expressed efficiently and purified effectively.The F1 antibody of one immune rabbit serum was positive for the three assays and the rank of antibody titer was ELISA ＞ GICA ＞ IHA(1 ∶ 10 × 211 vs 1 ∶ 10 × 29 vs 1 ∶ 10 × 2s).Among 1 685 field serum samples detected,89 serums(5.28％) werepositive for GICA,53(3.15％) were positive for IHA and 101 (5.99％) were positive for ELISA.General coincidence rates for GICA,IHA and ELISA were 97.3％ and 97.9％,respectively.Chi-square test for paired data showed that the positive rate of GICA was higher than that of IHA(x2 =26.63,P ＜ 0.05),but the difference between GICA and ELISA was not statistically significant(x2=3.36,P＞ 0.05).Conclusions rF1 protein is successful expressed and purified.The specificity and sensitivity of the GICA established using rF1 are higher than those of IHA and ELISA.When GICA is combined with IHA and ELISA,the time and quality of plague diagnosis may be improved effectively.
