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ABSTRACT: 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.
Lab on a Chip 07/2012; 12(20):3907-13. · 5.67 Impact Factor
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ABSTRACT: The application of microfluidic droplet PCR for single-molecule amplification and analysis has recently been extensively studied. Microfluidic droplet technology has the advantages of compartmentalizing reactions into discrete volumes, performing highly parallel reactions in monodisperse droplets, reducing cross-contamination between droplets, eliminating PCR bias and nonspecific amplification, as well as enabling fast amplification with rapid thermocycling. Here, we have reviewed the important technical breakthroughs of microfluidic droplet PCR in the past five years and their applications to single-molecule amplification and analysis, such as high-throughput screening, next generation DNA sequencing, and quantitative detection of rare mutations. Although the utilization of microfluidic droplet single-molecule PCR is still in the early stages, its great potential has already been demonstrated and will provide novel solutions to today's biomedical engineering challenges in single-molecule amplification and analysis.
Analytical and Bioanalytical Chemistry 03/2012; 403(8):2127-43. · 3.78 Impact Factor
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ABSTRACT: We have developed a microfluidic method to produce monodisperse, size-controllable silica microspheres via hydrolysis and polymerization of TEOS at the interface of water-in-oil droplets. By altering the concentration of TEOS in oil phase and CTAB in aqueous phase, we achieved control over morphology of the microspheres from hollow, to partly hollow, to solid. The hollow silica microsphere can be used for rapid waste removal and detoxification extraction with a very simple procedure.
ACS Applied Materials & Interfaces 09/2010; · 4.53 Impact Factor
