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Highly sensitive and quantitative detection of rare pathogens through agarose droplet microfluidic emulsion PCR at the single-cell level.
MedLine Citation:
PMID:  22836582     Owner:  NLM     Status:  Publisher    
Abstract/OtherAbstract:
Genetic alternations can serve as highly specific biomarkers to distinguish fatal bacteria or cancer cells from their normal counterparts. However, these mutations normally exist in very rare amount in the presence of a large excess of non-mutated analogs. Taking the notorious pathogen E. coli O157:H7 as the target analyte, we have developed an agarose droplet-based microfluidic ePCR method for highly sensitive, specific and quantitative detection of rare pathogens in the high background of normal bacteria. Massively parallel singleplex and multiplex PCR at the single-cell level in agarose droplets have been successfully established. Moreover, we challenged the system with rare pathogen detection and realized the sensitive and quantitative analysis of a single E. coli O157:H7 cell in the high background of 100 000 excess normal K12 cells. For the first time, we demonstrated rare pathogen detection through agarose droplet microfluidic ePCR. Such a multiplex single-cell agarose droplet amplification method enables ultra-high throughput and multi-parameter genetic analysis of large population of cells at the single-cell level to uncover the stochastic variations in biological systems.
Authors:
Zhi Zhu; Wenhua Zhang; Xuefei Leng; Mingxia Zhang; Zhichao Guan; Jiangquan Lu; Chaoyong James Yang
Publication Detail:
Type:  JOURNAL ARTICLE     Date:  2012-7-27
Journal Detail:
Title:  Lab on a chip     Volume:  -     ISSN:  1473-0189     ISO Abbreviation:  -     Publication Date:  2012 Jul 
Date Detail:
Created Date:  2012-7-27     Completed Date:  -     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  101128948     Medline TA:  Lab Chip     Country:  -    
Other Details:
Languages:  ENG     Pagination:  -     Citation Subset:  -    
Affiliation:
State Key Laboratory of Physical Chemistry of Solid Surfaces, Key Laboratory for Chemical Biology of Fujian Province, Key Laboratory of Analytical Science, Department of Chemical Biology, College of Chemistry and Chemical Engineering; Xiamen University, Xiamen, Fujian, China361005. cyyang@xmu.edu.cn.
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