Deng Ming Jun, Ji Xin Cheng, Xiao Xi Zhi, Sun Tao, Wu Zhen Xing, Zheng Xiao Long, Wang Qun, Zhu Lai Hua
J AOAC Int. 2014 Mar-Apr;97(2):561-6. doi: 10.5740/jaoacint.13-069.
Giardia lamblia cysts at low concentrations were detected in water samples using a highly sensitive immunological-PCR (IPCR) method. Magnetic gold particles were precoated with monoclonal anti-Giardia antibodies, and Giardia lamblia cysts ranging from 1 to 100 cysts were diluted in 500 microL of water. A biotinylated detection antibody bound to the G. lamblia cysts was then linked by a streptavidin bridge to biotinylated Giardia-reporter DNA. After extensive washing, reporter DNA was released by denaturation, transferred to PCR tubes, amplified, electrophoresed, and visualized. An optimized immuno-PCR method detected as little as five G. lamblia cysts. To further evaluate the specificity and the clinical application of this IPCR assay for G. lamblia cysts, Entamoeba histolytica and Cryptosporidium parvum were also collected and detected by IPCR. The data demonstrated that this monoclonal antibody-based IPCR method is particularly useful for analysis of environmental water samples in which the densities of G. lamblia cysts is very low, and provides a platform capable of rapid screening of samples from drinking water, wells, rivers, lakes, and recreational swimming pools for trace levels of G. lamblia cysts.
采用高灵敏度免疫PCR（IPCR）方法检测水样中低浓度的蓝氏贾第鞭毛虫包囊。用抗蓝氏贾第鞭毛虫单克隆抗体预包被磁性金颗粒，将1至100个蓝氏贾第鞭毛虫包囊稀释于500微升水中。然后，与蓝氏贾第鞭毛虫包囊结合的生物素化检测抗体通过链霉亲和素桥与生物素化的贾第鞭毛虫报告基因DNA相连。经过充分洗涤后，通过变性释放报告基因DNA，转移至PCR管中进行扩增、电泳和可视化分析。优化后的免疫PCR方法可检测低至5个蓝氏贾第鞭毛虫包囊。为进一步评估该IPCR检测法对蓝氏贾第鞭毛虫包囊的特异性及临床应用，还收集了溶组织内阿米巴和微小隐孢子虫，并采用IPCR进行检测。数据表明，这种基于单克隆抗体的IPCR方法对于分析蓝氏贾第鞭毛虫包囊密度极低的环境水样特别有用，并提供了一个能够快速筛查来自饮用水、水井、河流、湖泊和休闲游泳池的样品中痕量蓝氏贾第鞭毛虫包囊的平台。
