Publication detail

A new microfluidic approach for the one-step capture, amplification and label-free quantification of bacteria in raw milk

Year: 2017

Type of publication: článek v odborném periodiku

Name of source: Chemical Science

Page from-to: 1329-1336

Titles:

Language	Name	Abstract	Keywords
cze	Nová mikrofluidní metoda pro zachycování, amplifikaci a kvantifikaci bakterií v mléce	Práce popisuje mikrofluidní metodu pro specifické zachycení a detekci infekčních bakterií na základě imunorekognitivní a proliferační energie. Využívá se zde magnetické mikrofluidní lože, kde jsou magnetické a proudící síly vyrovnány tak, aby se udržely funkcionalizované superparamagnetické částice uvnitř komůrky čipu během průchodu vzorku. Zachycené buňky se pak kultivují in situ pomocí infuze živného média. Systém byl validován přímou jednostupňovou detekcí Salmonella Typhimurium v nezředěném surovém mléce bez předchozího ošetření. Růst bakterií indukuje expanzi fluidního lože, zejména kvůli objemu který zaujímají nově vytvořené bakterie. Tato expanze může být pozorována pouhým okem, což umožňuje jednoduchou detekci pouze několika bakterií za několik málo hodin. Tato nová technologie umožňuje rychlou, přenosu a automatizovanou detekci bakterií pro různé aplikace v potravinách, životním prostředí, ale i klinických vzorcích.	PCR amplifikace; rychlá detekce; identifikace; čip; fluidika; kmeny; systém
eng	A new microfluidic approach for the one-step capture, amplification and label-free quantification of bacteria in raw milk	A microfluidic method to specifically capture and detect infectious bacteria based on immunorecognition and proliferative power is presented. It involves a microscale fluidized bed in which magnetic and drag forces are balanced to retain antibody-functionalized superparamagnetic beads in a chamber during sample perfusion. Captured cells are then cultivated in situ by infusing nutritionally-rich medium. The system was validated by the direct one-step detection of Salmonella Typhimurium in undiluted unskimmed milk, without pre-treatment. The growth of bacteria induces an expansion of the fluidized bed, mainly due to the volume occupied by the newly formed bacteria. This expansion can be observed with the naked eye, providing simple low-cost detection of only a few bacteria and in a few hours. The time to expansion can also be measured with a low-cost camera, allowing quantitative detection down to 4 cfu (colony forming unit), with a dynamic range of 100 to 107 cfu ml−1 in 2 to 8 hours, depending on the initial concentration. This mode of operation is an equivalent of quantitative PCR, with which it shares a high dynamic range and outstanding sensitivity and specificity, operating at the live cell rather than DNA level. Specificity was demonstrated by controls performed in the presence of a 500× excess of non-pathogenic Lactococcus lactis. The system's versatility was demonstrated by its successful application to the detection and quantitation of Escherichia coli O157:H15 and Enterobacter cloacae. This new technology allows fast, low-cost, portable and automated bacteria detection for various applications in food, environment, security and clinics.	PCR amplification; rapid detection; identification; chip; fluidization; strains; system

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