中华检验医学杂志
中華檢驗醫學雜誌
중화검험의학잡지
CHINESE JOURNAL OF LABORATORY MEDICINE
2010年
7期
587-593
,共7页
吴文鹤%陈勇%蒋璐茜%崔晓阳%聂小博%张杰%吕建新
吳文鶴%陳勇%蔣璐茜%崔曉暘%聶小博%張傑%呂建新
오문학%진용%장로천%최효양%섭소박%장걸%려건신
肠致病性大肠杆菌%比色法%SELEX适体技术%生物传感技术
腸緻病性大腸桿菌%比色法%SELEX適體技術%生物傳感技術
장치병성대장간균%비색법%SELEX괄체기술%생물전감기술
Enteropathogenic Escherichia coli%Colorimetry%SELEX aptamer technique%Biosensing techniques
目的 结合纳米技术,构建一种可以快速比色检测肠致病性大肠埃希菌的适配体生物传感器.方法 对已有的适配体序列进行截短设计,将截短后的适配体通过酰胺化反应修饰到PDA纳米囊泡的表面,实现生物传感器的组装.利用适配体与LPS间特异性结合和结合后引起PDA纳米囊泡的颜色变化和比色响应值(CR)改变来定性或定量检测溶液中的肠致病性大肠埃希菌.并通过透射电子显微镜对PDA纳米囊泡与肠致病性大肠埃希菌之间的作用进行验证.结果 成功获得具有结合肠致病性大肠埃希菌功能的适配体截短序列;成功构建可以快速比色检测肠致病性大肠埃希菌的适配体生物传感器,并经透射电镜证实.其不需要特殊仪器,操作简便,检测时间短,仅需30 min;灵敏度较高,检出限为105 CFU/ml,线性范围为105～108CFU/ml;稳定性高,对106 CFU/ml菌液检测的相对标准偏差为6.08%;特异性强,对肠致病性大肠杆菌0111的检测的特异性为100%.结论 本研究成功构建了肠致病性大肠埃希菌适配体生物传感器,实现对肠致病性大肠埃希菌的快速比色检测.
目的 結閤納米技術,構建一種可以快速比色檢測腸緻病性大腸埃希菌的適配體生物傳感器.方法 對已有的適配體序列進行截短設計,將截短後的適配體通過酰胺化反應脩飾到PDA納米囊泡的錶麵,實現生物傳感器的組裝.利用適配體與LPS間特異性結閤和結閤後引起PDA納米囊泡的顏色變化和比色響應值(CR)改變來定性或定量檢測溶液中的腸緻病性大腸埃希菌.併通過透射電子顯微鏡對PDA納米囊泡與腸緻病性大腸埃希菌之間的作用進行驗證.結果 成功穫得具有結閤腸緻病性大腸埃希菌功能的適配體截短序列;成功構建可以快速比色檢測腸緻病性大腸埃希菌的適配體生物傳感器,併經透射電鏡證實.其不需要特殊儀器,操作簡便,檢測時間短,僅需30 min;靈敏度較高,檢齣限為105 CFU/ml,線性範圍為105～108CFU/ml;穩定性高,對106 CFU/ml菌液檢測的相對標準偏差為6.08%;特異性彊,對腸緻病性大腸桿菌0111的檢測的特異性為100%.結論 本研究成功構建瞭腸緻病性大腸埃希菌適配體生物傳感器,實現對腸緻病性大腸埃希菌的快速比色檢測.
목적 결합납미기술,구건일충가이쾌속비색검측장치병성대장애희균적괄배체생물전감기.방법 대이유적괄배체서렬진행절단설계,장절단후적괄배체통과선알화반응수식도PDA납미낭포적표면,실현생물전감기적조장.이용괄배체여LPS간특이성결합화결합후인기PDA납미낭포적안색변화화비색향응치(CR)개변래정성혹정량검측용액중적장치병성대장애희균.병통과투사전자현미경대PDA납미낭포여장치병성대장애희균지간적작용진행험증.결과 성공획득구유결합장치병성대장애희균공능적괄배체절단서렬;성공구건가이쾌속비색검측장치병성대장애희균적괄배체생물전감기,병경투사전경증실.기불수요특수의기,조작간편,검측시간단,부수30 min;령민도교고,검출한위105 CFU/ml,선성범위위105～108CFU/ml;은정성고,대106 CFU/ml균액검측적상대표준편차위6.08%;특이성강,대장치병성대장간균0111적검측적특이성위100%.결론 본연구성공구건료장치병성대장애희균괄배체생물전감기,실현대장치병성대장애희균적쾌속비색검측.
Objective To develop and evaluate an aptamer based biosensor (aptasensor) for rapid colorimetric detection of enteropathogenic Escherichia coli (EPEC). Method The aptasensor was fabricated by modifying the truncated LPS-binding aptamer on the surface of nanoscale polydiacetylene vesicles using peptide bonding between the carboxyl group of the vesicle and the amine group of the aptamer. Molecular recognition between EPEC and aptamer at the interface of the vesicle led to blue-red transition of polydiacetylene which was readily visible to the naked eyes and could be quantified by colorimetric responses (CR). Transmission electron microscopy (TEM) was used to confirm the specific interactions between EPEC and polydiacetylene vesicles. Result Truncated aptamer showed the similar LPS-binding activity. The aptasensor could detect the target bacteria in a range of 105-108 colony-forming units (CFU)/ml within less than 30 minutes and its specificity was 100% for detection of EPEC O111. The sensor reproducibiliry obtained at 106 CFU/ml was 6. 08% R. S. D. The results of TEM confirmed that the specific interactions between EPEC and polydiacetylene vesicles. Conclusion A new aptasensor was developed successfully for rapid colorimetric detection of EPEC.
