[1]郭小兵△,代志峰,宋海容,等.CTXM38型超广谱β内酰胺酶快速检测方法的建立*[J].郑州大学学报(医学版),2014,(01):66.
 GUO Xiaobing,DAI Zhifeng,SONG Hairong,et al.Establishment of a quick detection method of CTXM38 type ESBLs[J].JOURNAL OF ZHENGZHOU UNIVERSITY(MEDICAL SCIENCES),2014,(01):66.
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CTXM38型超广谱β内酰胺酶快速检测方法的建立*
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《郑州大学学报(医学版)》[ISSN:1671-6825/CN:41-1340/R]

卷:
期数:
2014年01期
页码:
66
栏目:
应用研究
出版日期:
2014-01-20

文章信息/Info

Title:
Establishment of a quick detection method of CTXM38 type ESBLs
作者:
郭小兵代志峰宋海容张傅山叶亚菲明亮
郑州大学第一附属医院检验科 郑州 450052
Author(s):
GUO XiaobingDAI ZhifengSONG HairongZHANG FushanYE YafeiMING Liang
Clinical Laboratory, the First Affiliated Hospital, Zhengzhou University, Zhengzhou 450052
关键词:
CTXM38超广谱β内酰胺酶酶联免疫吸附实验
Keywords:
CTXM38ESBLsELISA
分类号:
R392
摘要:
目的:建立一种CTXM38型超广谱β内酰胺酶(ESBLs)的快速检测方法。方法:采用梅花形双向琼脂扩散实验检测酶蛋白与抗体反应最适比;根据双抗体夹心法原理设计酶联免疫吸附实验(ELISA)检测试剂盒;平行测定30次BL21pET28aCTXM38转化子表达产物,检测批内变异;分别采用所建立的ELISA法与PCR技术对146株产ESBLs大肠埃希菌进行检测,评价方法特异性;对27株非产CTXM38型产酶大肠埃希菌进行检测,计算假阳性率;对产CTXM38型ESBLs菌株不同时间段培养液进行检测,明确最适检测时间。结果:抗体与4 h培养物反应最适比为164;批内检测结果变异系数为1.525%;PCR及所建立的ELISA法检测产CTXM38型ESBLs菌株的检出率分别为4.11%和7.53%(χ2=2.286,P=0.131);交叉反应中存在假阳性现象(1/27);产酶菌株培养4 h后进行ESBLs检测较为合适。结论:所建立的方法适用于临床产CTXM38型ESBLs菌株的检测。
Abstract:
Aim: To establish a quick detection method of CTXM38 type ESBLs. Methods: The reaction optimal ratio between CTXM38 type ESBLs and polyclonal antibody was confirmed by double agar diffusion test. The ELISA kit was designed according to doubleantibody sandwich principle. The products of CTXM38 transformant were detected 30 times to obtain the information of experimental repeatability and intraassay variation;146 ESBLsproducing E.coli strains were detected by PCR and the designed method respectively to evaluate the experimental specificity; 27 isolates of ESBLsproducing E.coli strains(not carrying CTXM38 type ESBLs) were detected to evaluate the experimental crossreactivity;the culture medium of CTXM38 type ESBLsproducing E.coli was detected at various time to confirm the optimal detective time of ESBLs. Results: The reaction optimal ratio between CTXM38 type ESBLs and polyclonal antibody was 164; the results of the parallel testing indicated that the CV was 1.525%;the detection rate of PCR and ELISA was 4.11% and 7.53%(χ2=2.286,P=0.131); the designed ELISA might have the phenomenon of falsepositivity(1/27); the optimal detection time for ESBLs was being cultured for 4 h. Conclusion: The established ELISA can meet the need for detecting CTXM38type ESBLsproducing strains.

参考文献/References:

[1]Shu JC,Chia JH,Kuo AJ,et al. A 7year surveillance for ESBLproducing Escherichia coli and Klebsiella pneumoniae at a university hospital in Taiwan: the increase of CTXM15 in the ICU[J]. Epidemiol Infect,2010,138(2):253 [2]冯羡菊,苟建军,曾利,等.产超广谱β内酰胺酶肺炎克雷伯菌耐药监测及基因型分析[J].郑州大学学报:医学版,2009,44(5):1086 [3]郭小兵,代志峰,宋海容,等.CTXM38型超广谱β内酰胺酶抗体的制备及鉴定[J].山东医药,2012,52(19):29 [4]朱立平,陈学清.免疫学常用实验方法[M].北京:人民军医出版社,2000:78 [5]叶应妩,王毓三,申子瑜.全国临床检验操作规程 [M]. 3版.南京:东南大学出版社,2006. [6]徐灵彬,刘原,王香玲.西安地区肠杆菌科细菌的超广谱β内酰胺酶基因型的研究[J].西安交通大学学报:医学版,2008,29(4):443 [7]徐敏超,王晶,严群,等.宽噬噬菌体治疗实验性小鼠产超广谱β内酰胺酶大肠埃希菌感染的研究[J].中华医院感染学杂志,2006,16(3):252 [8]张志坚,阎志勇,张钦宪.郑州地区大肠埃希菌产超广谱β内酰胺酶基因型分布[J].郑州大学学报:医学版,2010,45(5):730 [9]Yu WL,Chuang YC,WaltherRasmussen J.Extendedspectrum betalactamases in Taiwan: epidemiology, detection, treatment and infection control[J].J Microbiol Immunol Infect,2006,39(4):264 [10]郭小兵,张志坚,阎志勇,等.CTXM14型超广谱β内酰胺酶原核表达载体的构建及其抗药活性分析[J].山东医药,2009,49(11):21 [11]张志坚,郭小兵,阎志勇,等.郑州地区大肠埃希菌中CTXM型超广谱β内酰胺酶的基因型分布[J].中华医院感染学杂志,2009,19(18):2382 [12]杜艳霞.酶联免疫吸附试验测定操作的体会[J].中国实用医药,2009,4(27):139 [13]孙桂梅,韩金玲,赵莉,等.免疫诊断(主要是ELISA)试验操作的几点体会[J].中国现代药物应用,2009,3(3):161

备注/Memo

备注/Memo:
*河南省医学科技攻关项目2011020014;△男,1971年8月生,博士,副教授,研究方向:细菌耐药机制,Email:gxbing928@126.com
更新日期/Last Update: 2014-02-20