摘要
为提高畜禽类食品中空肠弯曲菌的检出率和灵敏度 ,应用抗血清和磁性微珠首次制备弯曲菌免疫磁珠 ,利用弯曲菌免疫磁珠直接捕获检样中目的菌 ,不需要增菌培养 ;通过荧光PCR检测鞭毛蛋白A(flaA)基因和 或马尿酸酶 (hipO)基因 ,首次建立空肠弯曲菌的磁捕获_荧光聚合酶链反应 (IMC_FPCR)方法。IMC_FPCR法检测空肠弯曲菌方法简便易行 ,可在 2 4h内完成 ,特异性好 ,检测低限达到 10cfu mL ,抗干扰性强。IMC_FPCR方法可望解决非可培养状态的空肠弯曲菌检测难题 ,是一种适用于检验检疫、卫生防疫和农产品安全检验等领域的快速方法。
In order to develop a rapid method which can check Campylobacter jejuni in animal and poultry foods nicely, an immun omag netic capture-fluorescent PCR (IMC-FPCR) method was established in this paper. T he reported method involves isolation of the target pathogen by immunocapture pr ior to the fluorescent PCR step, therefore the immunomagnetic-beads for Campylob acter were developed, and two groups of primer/probe, which targeted for the spe cies special sequence of fl gene and hipO gene for Campylobacter jejuni were d esigned. The immunomagnetic capture-fluorescent PCR assay amplification of the h ipO gene and fl gene for detection of Campylobacter jejuni was firstly reporte d in this paper. Result indicated that IMC-FPCR method permits direct detection of the pathogen without an enrichment step and can be performed in approximately 24 h. The assay results are positive for all of the isolates of Campylobacter jejuni (3 isolates, including type strain ATCC 33560 and ATCC8341) with a det ecti on limit of approximately 10 cfu/mL, are negative for Campylobacter coli and sev eral other bacteria. IMC-FPCR assay provide not only a rapid, sensitive method f or quantitative detection of Campylobacter jejuni, but also an important met hod for detecting of Campylobacter jejuni of viable but non-culturerable (VN C) state.
出处
《生物工程学报》
CAS
CSCD
北大核心
2005年第2期336-340,共5页
Chinese Journal of Biotechnology
基金
国家质量监督检验检疫总局重大科技项目 (No .2 0 0 2IK0 0 2_0 6)~~
关键词
空肠弯曲菌
免疫磁珠捕获
荧光PCR
检测
Campylobacter jejuni, immunomagnetic ca pture, fluorescent PCR, detection
