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Průměrný učitel vypráví. Dobrý učitel vysvětluje. Výborný učitel ukazuje. Nejlepší učitel inspiruje.

Charles Farrar Browne

mendel

Výzkum

3D Printed Chip for Electrochemical Detection of Influenza Virus Labelled with CdS Quantum Dots

Influenza is likely the most powerful member of the group of potential pandemic agents (Krasnoselsky and Katze 2012), because of the continuous mutational changes in its surface antigens, hemagglutinin (HA) and neuraminidase (NA), which play the main role in the mechanism of their interaction with sialic acid (SA) receptors on a host cell (Chen et al. 2013). Particularly, HA is a trimeric glycoprotein expressed on the membrane of influenza virus (Suenaga et al. 2012), which binds to SA receptors on the surface of the host cell and subsequently mediates fusion of the viral and target membranes (McCullough et al. 2012). Tropism and adaptation of influenza viruses to new hosts depend partly on the distribution of the sialic acid receptors (Rudrawar et al. 2013). The most widely described mechanism indicates that human influenza virus prefers the SA-?-2,6-Gal terminal glycan, whereas the avian influenza virus prefer SA-?-2,3-Gal one (Chen et al. 2012). In addition, HA is considered to be the main target for antibodies upon vaccination as well as native infection (Lingwood et al. 2012; Shembekar et al. 2013). Vaccination is the most effective prophylactic method against influenza (Wang et al. 2012). Quantitative and qualitative analysis of vaccine antigens is a turning point before the vaccine is placed on the market and used for immunization (Williams et al. 2012).

Numerous analytical methods are used to detect influenza viruses including methods based on the direct isolation of viruses (Bui et al. 2013; El-Zoghby et al. 2012) followed by real time-polymerase chain reaction (RT-PCR) (Charrel and Salez 2013; Piralla et al. 2013; Tong et al. 2012), or immunology tests (Hemmatzadeh et al. 2013; Shembekar et al. 2013; Wang et al. 2012; Zhao et al. 2011). Most of them have some disadvantages (an isolation of virus is highly sensitive, but time consuming and laborious method; RT-PCR requires appropriate equipment and well-trained staff). For this reason, new sensors and biosensors based on magnetic beads separation are coming to the forefront (Kamikawa et al. 2010; Li et al. 2011; Lien et al. 2011). Its sensitivity, specificity, speed, and the ability to recognize a very low concentration of target molecules belong to the most important features of a biosensor (Krejcova et al. 2012b; Xu and Wang 2012). For this reason, wide range of methods, materials and substances to optimize developing biosensing instrument has been tested (Kim et al. 2013). Paramagnetic particles (MPs) are the excellent tool with many advantageous features, such as easy handling and possibility to be separated by the magnetic field. MPs have surface that can be modified easily to allow covalent binding or simple unspecific adsorption of biomolecules (proteins, antibodies, nucleic acids) (Gijs 2004; Ramadan and Gijs 2012). In addition, large surface area related to the low volume is another great advantage. Because of the mentioned reasons, these particles have found many applications to enhance the selectivity, sensitivity, and speed of isolation methods (Krejcova et al. 2012a). Currently, various bead-based assays implemented to microfluidic systems have been reported for biomedical applications (Kirby et al. 2012; Lien et al. 2011; Tsai et al. 2013; Zitka et al. 2011). Combination of advanced biological detection methods (e.g. nucleic acid-based or immunology-based protocols) with microfluidic and immunomagnetic separation techniques exploiting functionalized magnetic particles has tremendous potential for realization of an integrated system for detection of pathogens, in particular, of waterborne pathogens (Casavant et al. 2013; Grutzkau and Radbruch 2010; Ramadan and Gijs 2012; Yang et al. 2013; Yulita et al. 2010). The isolated nucleic acid can be detected directly or labelled in these systems. Quantum dots (QDs) having unique optic properties, such as high quantum yield, broad absorption spectra, narrow emission spectra, low bleaching, and excellent photo-stability, belong to such labels. They have found many applications in biomedical imaging (Liu et al. 2012; Semonin et al. 2012; Zhao et al. 2012), and also as electrochemically active labels (Krejcova et al. 2012b; Sobrova et al. 2013a; Sobrova et al. 2013b). In the current study, the microfluidic assay based on MPs based isolation of HA followed with the indirect detection of the isolated compound with QDs labels. Microfluidic device for isolation and detection of HA-QDs were fabricated using three-dimensional (3D) printing, which is an example of additive manufacturing or of solid freeform fabrication technology (Polzin et al. 2013). Unlike traditional fabrication, the additive fabrication refers adding layers instead of cutting them off. This technique has been used for decades for the "rapid production of prototypes" (Leigh et al. 2012; Pawlyn 2013). A big step forward in 3D printing in the last decade has been made, therefore the limits of 3D printing are continually pushing out (Pawlyn 2013).

Podpořeno projekty: CEITEC CZ.1.05/1.1.00/02.0068

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