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Cell-based biosensors appear to be an attractive tool for the rapid, simple, and cheap monitoring of chemotherapy effects at a very early stage. In this study, electrochemical measurements using a four-point probe method were evaluated for suspensions of four cancer cell lines of different tissue origins: SK–N–SH, HeLa, MCF-7 and MDA-MB-231, all for two different population densities: 50 K and 100 K cells/500 μL. The anticancer agent doxorubicin was applied for each cell type in order to investigate whether the proposed technique was able to determine specific differences in cell responses before and after drug treatment. The proposed methodology can offer valuable insight into the frequency-dependent bioelectrical responses of various cellular systems using a low frequency range and without necessitating lengthy cell culture treatment. The further development of this biosensor assembly with the integration of specially designed cell/electronic interfaces can lead to novel diagnostic biosensors and therapeutic bioelectronics.

European mistletoe (Viscum album) is a medicinal plant with significant anticancer properties. In vitro callus production provides an essential alternative for biomass production and medicinal compounds propagation. In this study, we investigated (a) the biotechnological production of somaclonal proteins from mistletoe callus propagation and (b) the in vitro cytotoxic properties of these mistletoe protein extracts against cervical cancer. Mistletoe proteins (MPEs) were extracted from natural resources (plant leaves and stems) and in vitro propagated variant calluses. Potential cytotoxic effects of the extracted proteins on HeLa cells were assessed by the MTT proliferation assay, whereas oxidative damage and induction of apoptosis were measured fluorometrically by the reactive oxygen species (ROS) sensitive carboxy-H2-DCFDA method and caspase-3 activation assays, respectively. Electrophoretic results indicated differences in the protein bands between callus and donor plant tissues. Leaf and stem protein extracts (PEs) demonstrated a strong impact on HeLa cells viability by promoting oxidative damage and inducing apoptosis in contrast to callus PEs. This study shows that MPEs application in HeLa cells causes oxidative alterations and apoptosis through caspase-3 activation and increased intracellular ROS levels. These findings may be useful for uncovering novel bioactive products of pharmaceutical interest as well as for gaining an understanding of the biochemical pathways of mistletoe proteins in cancer treatment. In spite of the low cytotoxicity observed for biotechnologically propagated somaclonal variant calluses, towards cervical cancer, the ability to establish biotechnological production of said mistletoe extracts is important because it allows for standardization of protein production.

One of the key challenges of the recent COVID-19 pandemic is the ability to accurately estimate the number of infected individuals, particularly asymptomatic and/or early-stage patients. We herewith report the proof-of-concept development of a biosensor able to detect the SARS-CoV-2 S1 spike protein expressed on the surface of the virus. The biosensor is based on membrane-engineered mammalian cells bearing the human chimeric spike S1 antibody. We demonstrate that the attachment of the protein to the membrane-bound antibodies resulted in a selective and considerable change in the cellular bioelectric properties measured by means of a Bioelectric Recognition Assay. The novel biosensor provided results in an ultra-rapid manner (3 min), with a detection limit of 1 fg/mL and a semi-linear range of response between 10 fg and 1 μg/mL. In addition, no cross-reactivity was observed against the SARS-CoV-2 nucleocapsid protein. Furthermore, the biosensor was configured as a ready-to-use platform, including a portable read-out device operated via smartphone/tablet. In this way, we demonstrate that the novel biosensor can be potentially applied for the mass screening of SARS-CoV-2 surface antigens without prior sample processing, therefore offering a possible solution for the timely monitoring and eventual control of the global coronavirus pandemic.

The development of methods for the detection of aflatoxin B1 (AFB1) in foods is a very important practice for ensuring food quality and safety. Most tests of AFB1 are still conducted with conventional methods (i.e. antibody-based ELISA tests, high performance liquid chromatography – HPLC); however biosensor methods are being developed to date as screening tools for field analysis. Compared to immunology/ELISA-like tests or chromatography methods, biosensors are able to provide rapid, sensitive, robust and cost effective quantitative methods for on-site testing. In this work we propose a cellular biosensor based on Vero cells, membrane engineered with anti-AFB1 antibody as the biological recognition element reacting with AFB1 molecules on gold nanoparticle/screen printed electrodes (SPEs) (three electrode system). In order to culture the cells on the SPEs surfaces the working electrodes were coated with poly- l -lysine to facilitate cell adhesion. The SPEs were connected to a potentiostat device through a transducer and chronoamperometric (CA) and cyclic voltammetric (CV) measurements were performed. Quantitative results obtained using the cellular biosensor method for AFB1 were compared to those obtained using the HPLC method in pistachio samples spiked with AFB1. The method displayed good sensitivity (r2 = 0.87) and detection limit (0.5 ng/mL).

This study presents a bioelectric cell-based biosensor for the monitoring of the pyrethroid pesticide cypermethrin, a voltage-gated sodium channel blocker, in tobacco samples. For this purpose, neuroblastoma cells were used as biorecognition elements. The potential interference by the tobacco major alkaloid nicotine on the detection of cypermethrin was also studied. In addition, fluorescence microscopy revealed a specific pattern of neuroblastoma cell calcium efflux (Ca2+) after treatment with nicotine or cypermethrin. Finally, actual field-derived tobacco extracts were used for assessing matrix effects on the biosensor&rsquo, s performance. The biosensor could detect cypermethrin in concentrations up to 1.5 &mu, g mL&minus, 1 without being influenced by the presence of nicotine and possibly other tobacco alkaloids. Though not selective for cypermethrin, the neuroblastoma-based biosensor system appears to be a promising alternative to laborious analysis methodologies for rapid, high throughput and cost-efficient screening of this pyrethroid in tobacco samples in the near future.

Background: In vitro cell culture monitoring can be used as an indicator of cellular oxidative stress for the assessment of different chemotherapy agents. Methods: A cell-based bioelectric biosensor was used to detect alterations in superoxide levels in the culture medium of HeLa cervical cancer cells after treatment with the chemotherapeutic agent 5-fluorouracil (5-FU). The cytotoxic effects of 5-fluorouracil on HeLa cells were assessed by the MTT proliferation assay, whereas oxidative damage and induction of apoptosis were measured fluorometrically by the mitochondria-targeted MitoSOX™ Red and caspase-3 activation assays, respectively. Results: The results of this study indicate that 5-FU differentially affects superoxide production and caspase-3 activation when applied in cytotoxic concentrations against HeLa cells, while superoxide accumulation is in accordance with mitochondrial superoxide levels. Our findings suggest that changes in superoxide concentration could be detected with the biosensor in a non-invasive and rapid manner, thus allowing a reliable estimation of oxidative damage due to cell apoptosis. Conclusions: These findings may be useful for facilitating future high throughput screening of different chemotherapeutic drugs with a cytotoxic principle based on free radical production.

Plant micropropagation has been proposed as a promising solution for the improvement of medicinal plants and the commercial production of phytochemicals. Tree Wormwood (Artemisia arborescens, Asteraceae) has been identified as a promising source for chamazulene production, a phytochemical compound with anti-inflammatory activity. However, the concurrent presence of β-thujone, an acute toxic agent is a significant limitation of Tree Wormwood utilization in essential oil (EO) production. Present study is aiming to develop in vitro cultivation protocols of tree wormwood and study the effects of plant growth regulators (PGRs) and culture conditions on the callus production of volatile metabolites (VM) and its equilibrium of chamazulene and β-thujone. Three treatments all containing the auxin 1-Naphthaleneacetic acid (NAA), and each one respectively the cytokines Kinetin (K), 16-Benzylaminopurine (BN), and Thidiazuron (TD) in Murashige and Skoog (MS) basal media, were applied under constant dark (D), while K and BN were also grown under light (L). Both EO and VMs, retrieved from the naturally and laboratory grown biomass were analyzed by a Gas Chromatographer-Mass Spectrometer (GC–MS) coupled with Gas Chromatographer-Flame Ionization Detector (GC-FID). The biotechnologically produced EOs presented a significantly increased yield per area and time, while callogenesis PGRs combinations and culture conditions exhibited promising results on the induction of chamazulene biosynthesis.

We now have diverse types of nanomaterials (NMs) comprising of inorganic particles (e.g., oxides, metals, and salts existing in nature or produced in the laboratory) or organic particles (e.g., polymer-clay nanocomposites and quantum dots that may be manufactured only in the laboratory), ranging in dimensions between 1 and 100 nm. Their unique physicochemical properties are determined by their shape, size, surface area, and charge. Because of the widespread application of NMs in various fields, an intentional or unintended release of nanomaterials into the environment is on the increase, while their adverse effects on biological systems are difficult to predict. This situation necessitates the assessment of the potential effects of NMs on the exposed organisms and ecological processes. This chapter summarizes the latest research findings regarding the impact of NMs on the growth, metabolism, and toxicity of nonvascular plants.

Cancer cell lines are important tools for anticancer drug research and assessment. Impedance measurements can provide valuable information about cell viability in real time. This work presents the proof-of-concept development of a bioelectrical, impedance-based analysis technique applied to four adherent mammalian cancer cells lines immobilized in a three-dimensional (3D) calcium alginate hydrogel matrix, thus mimicking in vivo tissue conditions. Cells were treated with cytostatic agent5-fluoruracil (5-FU). The cell lines used in this study were SK-N-SH, HEK293, HeLa, and MCF-7. For each cell culture, three cell population densities were chosen (50,000, 100,000, and 200,000 cells/100 &mu, L). The aim of this study was the extraction of mean impedance values at various frequencies for the assessment of the different behavior of various cancer cells when 5-FU was applied. For comparison purposes, impedance measurements were implemented on untreated immobilized cell lines. The results demonstrated not only the dependence of each cell line impedance value on the frequency, but also the relation of the impedance level to the cell population density for every individual cell line. By establishing a cell line-specific bioelectrical behavior, it is possible to obtain a unique fingerprint for each cancer cell line reaction to a selected anticancer agent.

(1) Background: Fungal metabolites such as haloanisoles (especially 2,4,6-tribromoanisole/ 2,4,6-TCA) are contaminants of cork and wood barrels, materials that are widely used in the wine industry. Thus, the accurate and timely detection of these substances is very important for this sector of beverage industry. (2) Methods: Potentiometry was used for the Bioelectric Recognition Assay (BERA)-based experimental approach, where changes in the electric properties of the Vero cells modified with anti-TCA antibodies were recorded in response to the presence of 2,4,6-TCA in different concentrations. Furthermore, a second electrochemical biosensor system based on the cyclic voltammetric (CV) measurement of a reaction taking place on a screen printed electrode was developed in parallel to the customized application and configuration of the cell-based system. (3) Results: The BERA cell-based biosensor was able to quantitatively differentiate among the lower 2,4,6-TCA concentrations (control, 0.25 and 1.25 ng/L) from spiked oak barrel water extracts in an entirely distinct and reproducible manner. In contrast, the CV method was not sensitive enough to differentiate between the samples. (4) Conclusions: The present study indicates that the BERA-based biosensor after further development and optimization could be used for the routine, high throughput detection of 2,4,6-TCA in oak barrel water extracts.

Prostate-specific antigen (PSA) is the established routine screening tool for the detection of early-stage prostate cancer. Given the laboratory-centric nature of the process, the development of a portable, ultra rapid high-throughput system for PSA screening is highly desirable. In this study, an advancedpoint-of-care system for PSA detection in human serum was developed based on a cellular biosensor where the cell membrane was modified by electroinserting a specific antibody against PSA. Thirty nine human serum samples were used for validation of this biosensory system for PSA detection. Samples were analyzed in parallel with a standard immunoradiometric assay (IRMA) and an established electrochemical immunoassay was used for comparison purposes. They were classified in three different PSA concentration ranges (0, &lt, 4 and &ge, 4 ng/mL). Cells membrane-engineered with 0.25 &mu, g/mL anti-PSA antibody demonstrated a statistically lower response against the upper (&ge, 4 ng/mL) PSA concentration range. In addition, the cell-based biosensor performed better than the immunosensor in terms of sensitivity and resolution against positive samples containing &lt, 4 ng/mL PSA. In spite of its preliminary, proof-of-concept stage of development, the cell-based biosensor could be used as aninitiative for the development of a fast, low-cost, and high-throughput POC screening system for PSA.

Mycotoxins and haloanisoles are secondary metabolites produced under special conditions of temperature and humidity by fungi colonizing a variety of commodities from preharvest up to consumer use. Ochratoxin A and 2,4,6-trichloanisole are produced mainly by species of the genus Aspergillus and Penicillium. Ochratoxin A exhibits nephrotic effects and can, potentially, be associated with human carcinogenesis, whereas 2,4,6-trichloanisole is primarily responsible for cork taint in wines. This review provides an overview of recent advances in biosensor technology for the determination of the aforementioned compounds in wine, beer and other beverages, as well as cork stoppers, which help in establishing and carrying out proper product quality-management strategies. Such a detailed investigation of biosensor-based detection methods of these toxic compounds in beverages could lead to the provision of safe-to-consume products, and allow the prioritization of future research efforts.

Two of the most important categories of pesticides used in agricultural practice are organophosphates and dithiocarbamates. Their extensive and inappropriate use has rendered their reliable monitoring at trace levels more and more necessary. This study presents the construction of a rapid and sensitive cellular biosensor test based on the measurement of changes of the cell membrane potential of immobilized cells, according to the working principle of the Bioelectric Recognition Assay (BERA). The cells were immobilized by entrapment in a sodium alginate bead and directly applied in different pesticide dilutions and agricultural samples. The pesticides used were the organophosphate insecticide diazinon and the dithiocarbamate fungicide propineb. Two different cell types, N2a (neuroblastoma) and Vero (fibroblast) were used as the biosensory elements in order to investigate their differential response against the pesticides. In this way, we hoped to increase the selectivity of the assay. Based on the observed patterns of response, we demonstrate that the sensor can be used for the qualitative and, in some concentrations, quantitative detection of the pesticides with a high degree of reproducibility. The lowest detected concentration was 3nM. Finally, for the investigation of the effects of different pesticides on the accumulation of cytosolic Ca(2+), we conducted a fluorescent assay on N2a cells treated with tomato sample extracts, which were replicates of the E.U. proficiency test sample. The tomato samples were either organically grown or contained 14 different pesticides. The experimental results showed a higher increase of the intracellular Ca(2+) concentration in cells treated with non-organic samples compared to the cells treated with organic samples.

The conventional analysis of pesticide residues in analytical commodities, such as tobacco and tobacco products is a labor intensive procedure, since it is necessary to cover a wide range of different chemicals, using a single procedure. Standard analysis methods include extensive sample pretreatment (with solvent extraction and partitioning phases) and determination by GC and HPLC to achieve the necessary selectivity and sensitivity for the different classes of compounds under detection. As a consequence, current methods of analysis provide a limited sample capacity. In the present study, we report on the development of a novel cell biosensor for detecting organophosphate and carbamate pesticide residues in tobacco. The sensor is based on neuroblastoma N2a cells and the measurement of changes of the cell membrane potential, according to the working principle of the Bioelectric Recognition Assay (BERA). The presence of pesticide residues is detected by the degree of inhibition of acetylcholine esterase (AChE). The sensor instantly responded to both the organophoshate pesticide chlorpyriphos and the carbamate carbaryl in a concentration-dependent pattern, being able to detect one part per billion (1 ppb). Additionally, tobacco leaf samples (in blended dry form) were analyzed with both the novel biosensor and conventional methods, according to a double-blind protocol. Pesticide residues in tobacco samples caused a considerable cell membrane hyperpolarization to neuroblastoma cells immobilized in the sensor, as indicated by the increase of the negative sensor potential, which was clearly distinguishable from the sensor’s response against pesticide-free control samples. The observed response was quite reproducible, with an average variation of +5,6%. Fluorescence microscopy observations showed that treatment of the cells with either chlorpyrifos or carbaryl was associated with increased [Ca2+]cyt . The novel biosensor offers fresh perspectives for ultra-rapid, sensitive and low-cost monitoring of pesticide residues in tobacco as well as other food and agricultural commodities.