Quartz Crystal Microbalance
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Procalcitonin is a blood protein and precursor of the hormone calcitonin. The procalcitonin level increases due to bacterial infections, sepsis, and other related pathologies. Here, we present a simple biosensor for procalcitonin assay suitable for point-of-care tests as an alternative to the current laboratory methods. The biosensor was based on a QCM piezoelectric sensor and a conjugate of gold nanoparticles-antibodies conjugate. It was suitable for the procalcitonin assay in biological samples and fully correlated to the standard ELISA method, and it did not suffer false positive or negative results or interferences. The detection limit was equal to 37.8 ng/l and the quantification limit to 104 ng/l for a sample of 25 μl. The dynamic range of the assay was 37.8 ng/l to 30.0 μg/l. The practical relevance of the biosensor is expected considering the findings, and the possible application of the assay principle for the development of biosensors for other markers is inferred.
Interferon gamma (IFNγ) is a cytokine and an immunochemical marker that can be used for revealing of infectious diseases and especially for distinguishing of viral and some types of bacterial infections. Blood tests for IFNγ are typically based on immunoassays like Enzyme-Linked Immunosorbent Assay (ELISA). In this paper, a biosensor working on the principle of quartz crystal microbalance (QCM) was developed as an alternative to the standard analytical methods for IFNγ. The biosensor contained antibodies against IFNγ immobilized on QCM and also on gold nanoparticles. A sandwich containing QCM, gold nanoparticles and IFNγ was formed and formation of the sandwich caused decrease of oscillation frequency. The assay exerted limit of detection 5.7 pg/ml for a sample sized 50 μl and one measuring cycle was finished within 90 min. The assay by biosensor fully correlated to standard ELISA. In a conclusion, the biosensor appears to be a fully applicable analytical tool for a simple assay of IFNγ. Overall simplicity and no special requirement on staff and equipment are the major advantages of the here presented assay.
- MeSH
- biosenzitivní techniky * MeSH
- imunoanalýza MeSH
- interferon gama MeSH
- kovové nanočástice * MeSH
- křemen MeSH
- lidé MeSH
- mikrorovnovážné techniky křemenného krystalu MeSH
- zlato MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Hyaluronan (HA) is a major component of peri- and extra-cellular matrices and plays important roles in many biological processes such as cell adhesion, proliferation and migration. The abundance, size distribution and presentation of HA dictate its biological effects and are also useful indicators of pathologies and disease progression. Methods to assess the molecular mass of free-floating HA and other glycosaminoglycans (GAGs) are well established. In many biological and technological settings, however, GAGs are displayed on surfaces, and methods to obtain the size of surface-attached GAGs are lacking. Here, we present a method to size HA that is end-attached to surfaces. The method is based on the quartz crystal microbalance with dissipation monitoring (QCM-D) and exploits that the softness and thickness of films of grafted HA increase with HA size. These two quantities are sensitively reflected by the ratio of the dissipation shift (ΔD) and the negative frequency shift (- Δf) measured by QCM-D upon the formation of HA films. Using a series of size-defined HA preparations, ranging in size from ~ 2 kDa tetrasaccharides to ~ 1 MDa polysaccharides, we establish a monotonic yet non-linear standard curve of the ΔD/ - Δf ratio as a function of HA size, which reflects the distinct conformations adopted by grafted HA chains depending on their size and surface coverage. We demonstrate that the standard curve can be used to determine the mean size of HA, as well as other GAGs, such as chondroitin sulfate and heparan sulfate, of preparations of previously unknown size in the range from 1 to 500 kDa, with a resolution of better than 10%. For polydisperse samples, our analysis shows that the process of surface-grafting preferentially selects smaller GAG chains, and thus reduces the average size of GAGs that are immobilised on surfaces comparative to the original solution sample. Our results establish a quantitative method to size HA and other GAGs grafted on surfaces, and also highlight the importance of sizing GAGs directly on surfaces. The method should be useful for the development and quality control of GAG-based surface coatings in a wide range of research areas, from molecular interaction analysis to biomaterials coatings.
Adsorption properties of protein Papain at the solid|liquid (0.1 M KCl) interfaces of different hydrophobicity [highly oriented pyrolytic graphite (HOPG), bare gold, CH3, OH, and COOH-terminated self-assembled monolayers on gold] were studied by a combined quartz crystal microbalance and atomic force microscopy techniques. It was found that Papain forms an incomplete monolayer at hydrophobic interfaces (HOPG and CH3-terminated substrate), whereas on more hydrophilic ones, a complete monolayer formation was always observed with either the onset of the formation of a second layer (bare gold substrate) or adsorption in a multilayer fashion, possibly a bilayer formation (OH-terminated substrate). The surface concentration and compact monolayer film thickness was much lower on the COOH-terminated substrate compared to other surfaces studied. This result was explained by partial dissociation of the interfacial COOH groups leading to additional electrostatic interactions between the positively charged protein domains and negatively charged carboxylate anions, as well as to local pH changes promoting protein denaturation.
Tumor Necrosis Factor Alpha (TNFα) is an important marker of inflammatory processes in human body. In the current healthcare, determination of TNFα blood or plasma level is done by Enzyme Linked Immuno-Sorbent Assay (ELISA) as a primary choice method. Piezoelectric immunosensors are analytical platform recording affinity interactions on their surface. It is inferred that the immunosensors would be a functional alternative to the ELISA. In this study, antibody against TNFα was immobilized on Quartz Crystal Microbalance (QCM) sensor and the same was made on magnetic particles. Human TNFα was measured in a way of interaction with QCM surface and then the particles were applied. The assay exerted sufficient limit of detection equal to 1.62pg/ml and it also fully correlated with standard ELISA tests. No interference by interleukin 6 or albumin was observed. Long term stability of the immunosensors lasting for at least three months was found. The immunosensors appears to be readily for practical performance and it would be an alternative to the standard ELISA especially when diagnoses made in field, homecare conditions or conditions of small hospitals as an emergency test.
Custom immuno-magnetic devices are desirable tools for biomedical and biotechnological applications. Herein, surface active maghemite nanoparticles (SAMNs) are proposed as a versatile platform for developing tailored immuno-magnetic nano-carriers by simple wet reactions. Two examples for conjugating native and biotinylated antibodies were presented along with their successful applications in the recognition of specific foodborne pathogens. Nanoparticles were functionalized with rhodamine B isothiocyanate (RITC), leading to a fluorescent nano-conjugate, and used for binding anti-Campylobacter fetus antibodies (SAMN@RITC@Anti-Cf). The microorganism was selectively captured in the presence of two other Campylobacter species (C. jejuni and C. coli), as verified by PCR. Alternatively, SAMNs were modified with avidin, forming a biotin-specific magnetic nano-carrier and used for the immobilization of biotinylated anti-Listeria monocytogenes antibodies (SAMN@avidin@Anti-Lm). This immuno-magnetic carrier was integrated in piezoelectric quartz crystal microbalance (QCM) sensor for the detection of L. monocytogenes in milk, showing a detection limit of 3 bacterial cells. The present work presents a new category of customized immuno-magnetic nano-carriers as a competitive option for suiting specific applications. Graphical abstract ᅟ.
- MeSH
- adjuvancia imunologická chemie MeSH
- avidin chemie MeSH
- Listeria imunologie MeSH
- magnetismus * MeSH
- mikrorovnovážné techniky křemenného krystalu metody MeSH
- monoklonální protilátky chemie MeSH
- nanočástice chemie MeSH
- povrchové vlastnosti MeSH
- transmisní elektronová mikroskopie MeSH
- železité sloučeniny chemie MeSH
- Publikační typ
- časopisecké články MeSH
Label-free immunosensors are well suited for detection of microorganisms because of their fast response and reasonable sensitivity comparable to infection doses of common pathogens. Active (lever oscillator and frequency counter) and passive (impedance analyzer) modes of quartz crystal microbalance (QCM) were used and compared for rapid detection of three strains of E. coli. Different approaches for antibody immobilization were compared, the immobilization of reduced antibody using Sulfo-SMCC was most effective achieving the limit of detection (LOD) 8 × 104 CFU·mL-1 in 10 min. For the passive mode, software evaluating impedance characteristics in real-time was developed and used. Almost the same results were achieved using both active and passive modes confirming that the sensor properties are not limited by the frequency evaluation method but mainly by affinity of the antibody. Furthermore, reference measurements were done using surface plasmon resonance. Effect of condition of cells on signal was observed showing that cells ruptured by ultrasonication provided slightly higher signal changes than intact microbes.
- MeSH
- elektrická impedance MeSH
- Escherichia coli izolace a purifikace MeSH
- imobilizované proteiny metabolismus MeSH
- imunoanalýza MeSH
- kalibrace MeSH
- mikrorovnovážné techniky křemenného krystalu metody MeSH
- mikroskopie atomárních sil MeSH
- povrchová plasmonová rezonance MeSH
- protilátky metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Chlorinated hydrocarbons are very often used and are relatively dangerous substances from healthy risk point of view. While manipulating with them, mainly in large volumes, individual protective equipment (IPE) must be used in a protection position. Users are supposed to know the construction material breakthrough time especially in case of long-term usage of personal IPE and in the situation when contamination of them is real. Studying connections between a chemical compound structure and the structure of IPE characterised by barrier materials enables us to understand present body protective devices protection quality and gives us an option to choose barrier materials with targeted properties. In this article there are results of breakthrough time of isolating protection folio with a butyl rubber barrier layer in relation to chlorinated ethanes. This material is used for protection of specialists of both Fire Rescue Brigades and the Czech Armed Forces Chemical Corps. The PIEZOTEST device has been used for detection of permeated chemicals. The Quartz Crystal Microbalance (QCM) sensor is a part of PIEZOTEST device.
- Klíčová slova
- lag-time, butyl rubber, steady state permeation rate, breakthrough time,
- MeSH
- časové faktory MeSH
- chlorované uhlovodíky * chemie MeSH
- elastomery chemie MeSH
- ethan analogy a deriváty chemie MeSH
- ethylendichloridy chemie MeSH
- mikrorovnovážné techniky křemenného krystalu statistika a číselné údaje MeSH
- nebezpečné látky chemie MeSH
- ochranné oděvy * MeSH
- permeabilita * MeSH
- rozpustnost MeSH
- testování materiálů metody statistika a číselné údaje MeSH
- trichlorethany chemie MeSH
- Publikační typ
- práce podpořená grantem MeSH
This paper points to some aspects of possible usage of a QCM method within a set of toxic compounds permeated concentration based on a permeation curves rate and after underdone calibration of a measurement system. Presented assesses can be used not only in favour of the Czech Armed Forces Chemical Corps specialists but subsequently also in favour of personnel protection for those working in an area with toxic compounds appearance.
Combined experimental and computational studies of lipid membranes and liposomes, with the aim to attain mechanistic understanding, result in a synergy that makes possible the rational design of liposomal drug delivery system (LDS) based therapies. The LDS is the leading form of nanoscale drug delivery platform, an avenue in drug research, known as "nanomedicine", that holds the promise to transcend the current paradigm of drug development that has led to diminishing returns. Unfortunately this field of research has, so far, been far more successful in generating publications than new drug therapies. This partly results from the trial and error based methodologies used. We discuss experimental techniques capable of obtaining mechanistic insight into LDS structure and behavior. Insight obtained purely experimentally is, however, limited; computational modeling using molecular dynamics simulation can provide insight not otherwise available. We review computational research, that makes use of the multiscale modeling paradigm, simulating the phospholipid membrane with all atom resolution and the entire liposome with coarse grained models. We discuss in greater detail the computational modeling of liposome PEGylation. Overall, we wish to convey the power that lies in the combined use of experimental and computational methodologies; we hope to provide a roadmap for the rational design of LDS based therapies. Computational modeling is able to provide mechanistic insight that explains the context of experimental results and can also take the lead and inspire new directions for experimental research into LDS development. This article is part of a Special Issue entitled: Biosimulations edited by Ilpo Vattulainen and Tomasz Róg.
- MeSH
- lékové transportní systémy * MeSH
- lipidové dvojvrstvy chemie MeSH
- liposomy * MeSH
- membránové lipidy chemie MeSH
- mikrorovnovážné techniky křemenného krystalu MeSH
- polyethylenglykoly MeSH
- radiační rozptyl MeSH
- simulace molekulární dynamiky * MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- přehledy MeSH
