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Pathogenic bacteria detection using electrochemical immunosensors

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PASTUCHA Matěj FARKA Zdeněk JUŘÍK Tomáš MIKUŠOVÁ Zuzana SKLÁDAL Petr

Druh Článek ve sborníku
Konference XVIII. Workshop of Biophysical Chemists and Electrochemists
Fakulta / Pracoviště MU

Středoevropský technologický institut

Citace
WWW http://www.sci.muni.cz/~labifel/files/soubory/sbornik_2018.pdf#page=35
Klíčová slova Biosensor; Electrochemical Impedance Spectroscopy; Immunosensor; Label-free detection; Salmonella Typhimurium
Popis Pathogenic bacteria are a serious threat to human health and an important target for detection in key areas – public health, security, food safety and environment protection. With globalization, world is becoming more interconnected, people and goods are travelling further and faster than before. In such environment, there is a growing need for rapid, sensitive, selective and affordable techniques for microbial detection and identification. Traditional culturing methods are laborious and lengthy, usually taking days to provide results. More advanced methods based on molecular identification, most importantly polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA), are substantially faster, but require sophisticated laboratory equipment and trained personnel. Besides these well-established methods, biosensors are extensively developed as an alternative promising rapid and sensitive detection, suitable for screening and monitoring applications, on-the-spot measurement and providing ease of operation. In this project, a portable label-free impedimetric immunosensor for Salmonella Typhimurium detection was developed. The heart of the biosensor is a gold screen-printed electrode with immobilized specific antibody, which is incubated with sample. Bacterial cells present in the sample are specifically bound to surface of the electrode by the antibody and change of surface coverage is measured as a change in impedance. Several ways of sample processing (heat-treatment, sonication and their combination) were tested and their effect on the assay performance was evaluated. Impact of the sample treatment on cell shape and its binding onto the electrode was studied in detail by atomic force microscopy. The developed immunosensor allowed specific and sensitive detection of Salmonella Typhimurium with limit of detection of 10^3 CFU·mL-1, wide linear range up to 108 CFU·mL-1, negligible interference from other bacteria and short analysis time of 20 min. Suitability of the sensor for analysis of real samples was demonstrated by successful detection of Salmonella in milk. Furthermore, the developed immunosensor was combined with a home-built Arduino-based portable impedance analyzer to make on-the-spot measurements possible. No pre-enrichment of bacteria was utilized, which would bring the assay sensitivity to a much higher level, at the cost of longer analysis time. Sensitivity can be also improved by employing various nanoparticles or a labelled tracer antibody for signal enhancement by enzymatic precipitation. It is also possible to adapt the sensors for detection of other bacteria by replacing the specific antibodies.
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