Wang Yi, Li Hui, Wang Yan, Li Hua, Luo Lijuan, Xu Jianguo, Ye Changyun
State Key Laboratory of Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Chinese Center for Disease Control and Prevention, Changping, Beijing.
State Key Laboratory of Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Chinese Center for Disease Control and Prevention, Changping, Beijing; Department of Microbiology, GuiZhou Medical University, Guiyang, Guizhou, People's Republic of China.
Int J Nanomedicine. 2017 Jan 12;12:473-486. doi: 10.2147/IJN.S123625. eCollection 2017.
, one of most problematic foodborne pathogens, is responsible for listeriosis in both humans and animals and mainly transmitted through the food chain. In this report, we propose a simple, rapid, and nearly instrument-free molecular technique using multiple cross displacement amplification (MCDA) label-based gold nanoparticles lateral flow biosensor (LFB) for specific, sensitive, and visual detection of . The MCDA-LFB method was carried out at a constant temperature (61°C) for only 20 min during the reaction stage, and then the amplification mixtures were directly detected by using LFB, eliminating the use of an electrophoresis instrument, special reagents, or amplicon analysis equipment. The whole procedure, from sample processing to result indicating, was finished within 1 h. The analytical specificity of MCDA-LFB method was successfully determined by distinguishing the target bacterium from other pathogens. The analytical sensitivity of the MCDA-LFB assay was 10 fg of genomic templates per reaction in pure culture, which was in complete accordance with MCDA by gel electrophoresis, real-time turbidity, and colorimetric indicator. The assay was also successfully applied to detecting in pork samples. Therefore, the rapidity, simplicity, and nearly equipment-free platform of the MCDA-LFB technique make it possible for food control, clinical diagnosis, and more. The proof-of-concept assay can be reconfigured to detect various target sequences by redesigning the specific MCDA primers.
作为最具问题的食源性病原体之一,可导致人类和动物患李斯特菌病,主要通过食物链传播。在本报告中,我们提出了一种简单、快速且几乎无需仪器的分子技术,即使用基于多重交叉置换扩增(MCDA)标记的金纳米颗粒侧向流动生物传感器(LFB)来特异性、灵敏且可视化地检测[病原体名称未给出]。MCDA-LFB方法在反应阶段于恒温(61°C)下仅进行20分钟,然后使用LFB直接检测扩增混合物,无需使用电泳仪器、特殊试剂或扩增子分析设备。从样品处理到结果显示的整个过程在1小时内完成。通过将目标细菌与其他病原体区分开来,成功确定了MCDA-LFB方法的分析特异性。MCDA-LFB检测在纯培养物中的分析灵敏度为每个反应10 fg基因组模板,这与通过凝胶电泳、实时浊度和比色指示剂进行的MCDA完全一致。该检测方法还成功应用于猪肉样品中[病原体名称未给出]的检测。因此,MCDA-LFB技术的快速性、简单性和几乎无需设备的平台使其在食品控制、临床诊断等方面成为可能。通过重新设计特定的MCDA引物,可将概念验证检测重新配置以检测各种目标序列。