微流控免疫芯片富集捕获EHECO157︰H7的研究.docVIP

微流控免疫芯片富集/捕获EHEC O157︰H7的研究 管潇1，张惠静1，毕颖楠1，张莉1，郝敦玲1，韦劲宇2（第三军医大学：1医学检验系分析化学教研室，2学员旅3队，重庆 400038） 摘 要：目的　探讨基于免疫微珠的微流控芯片识别捕获EHEC O157︰H7的效率。方法　采用化学共价连接的方法将抗EHEC O157︰H7抗体固定于粒径为50μm的玻璃微珠上，再将微珠填充于PDMS/玻璃复合芯片中，建立由流体控制、免疫反应构成的细菌分选芯片；通过对进样流速、冲洗流速及冲洗时间的优化，在降低芯片对细菌的非特异性吸附的同时，增加芯片的特异性捕获效率。结果 在10μl·/min-1和5min的最佳冲洗流速和冲洗时间下，该芯片在10分钟内可完成对min内可完成对EHEC O157︰H7的捕获，捕获效率达到40%～63.96%，且实验组与对照组有显著差异且实验组与对照组有统计学差异（P0.01）。结论 本研究构建的EHEC O157︰H7识别芯片具有较高的捕获效率，为下一步构建灵敏、快速、微缩便携的细菌快速检测系统奠定了前期实验基础。 关键词：微流控芯片；免疫微珠；细菌捕获 中图法分类号：Q93-332；Q93-338 文献标识码：A A microfluidic chip forto bacteria concentrationing and captureing EHEC O157︰H7 GUAN Xiao, ZHANG Hui-jing, BI Ying-nan, ZHANG Li, HAO Dun-ling (Department of Analytical Chemistry, FacultyCollege of Clinical Laboratory SciencesMedicine, The Third Military Medical University, Chongqing, 400038, China) Abstract: Objective To study the efficiency of the concentration and capture of enterohemorrhagic Escherichia coli (EHEC) EHEC O157∶H7 by a self-made microfluidic chip based on microbead. Methods The anti-EHEC O157∶H7 antibody was first attached on the microbeads by direct covalent linkage，, then the microbeads were filled into microfluidic chip. After that, tThus, the bacteria capture chip was set up. Totally 1.6×109 CFU fluorescence labeled bacteria flowed into the chip at a sampling speed of 1, 2 and 5 μl/min respectively, and the elution speed of 0.5, 2, 5, 10 and 20 μl/min. The specific capture efficiency of the chip was strengthened through the optimization of sampling speed, elution speed and flow rate and rinsing time. Results The best sampling speed, elution speed and timerinsing flow rate and time were 5 μl/min, respectively 10 μLl·/min-1 and 5 min respectively. There was significant difference (P0.01) between the test group and the control group (P0.01). Conclusion Our self-made Bbacteria capture chip in this research got is of high catching efficiency, which had laidy a good base for the construction of rapid detection