Fiber-optic immunosensor for detection of Crimean-Congo Hemorrhagic fever IgG antibodies in patients

Crimean-Congo hemorrhagic fever (CCHF) is a severe viral disease with high fatality rate. CCHF virus is endemic in parts of Africa, Asia, Middle East and southeastern Europe. Rapid diagnostics of CCHF is vital for appropriate clinical management and prevention of secondary spread from human-to-human. Currently, diagnostics relies on Real-Time RT-PCR and antibody or antigen detection using ELISA. These methods require trained personnel and expensive equipment and are not appropriate for point-of-care (POC) diagnostics. Furthermore there are no POC assays available for CCHF. We developed a fiber-optic biosensor for the detection of CCHF IgG antibodies. In order to improve sensitivity, we optimized both the bioreceptor immobilization protocol and the chemiluminescence substrate formulation. The resulting protocol showed a 100-fold greater sensitivity for detection of CCHF antibodies. Finally, we evaluated the fiber-optic biosensor with two CCHF patient sera. We showed that the fiber-optic biosensor is 10-times more sensitive that colorimetric ELISA and is able to detect both patients with high and low levels of IgG antibodies. We believe that the fiber-optic biosensor is a suitable alternative to ELISA as it is much more sensitive and makes it possible to detect small amount of antibodies at an early stage of infection, and can be integrated as a point-of-care diagnostic system of CCHF

Cell-Based Sensor System Using L6 Cells for Broad Band Continuous Pollutant Monitoring in Aquatic Environments

Pollution of drinking water sources represents a continuously emerging problem in global environmental protection. Novel techniques for real-time monitoring of water quality, capable of the detection of unanticipated toxic and bioactive substances, are urgently needed. In this study, the applicability of a cell-based sensor system using selected eukaryotic cell lines for the detection of aquatic pollutants is shown. Readout parameters of the cells were the acidification (metabolism), oxygen consumption (respiration) and impedance (morphology) of the cells. A variety of potential cytotoxic classes of substances (heavy metals, pharmaceuticals, neurotoxins, waste water) was tested with monolayers of L6 cells (rat myoblasts). The cytotoxicity or cellular effects induced by inorganic ions (Ni2+ and Cu2+) can be detected with the metabolic parameters acidification and respiration down to 0.5 mg/L, whereas the detection limit for other substances like nicotine and acetaminophen are rather high, in the range of 0.1 mg/L and 100 mg/L. In a close to application model a real waste water sample shows detectable signals, indicating the existence of cytotoxic substances. The results support the paradigm change from single substance detection to the monitoring of overall toxicity

Are luminescent bacteria suitable for online detection and monitoring of toxic compounds in drinking water and its sources?

Biosensors based on luminescent bacteria may be valuable tools to monitor the chemical quality and safety of surface and drinking water. In this review, an overview is presented of the recombinant strains available that harbour the bacterial luciferase genes luxCDABE, and which may be used in an online biosensor for water quality monitoring. Many bacterial strains have been described for the detection of a broad range of toxicity parameters, including DNA damage, protein damage, membrane damage, oxidative stress, organic pollutants, and heavy metals. Most lux strains have sensitivities with detection limits ranging from milligrams per litre to micrograms per litre, usually with higher sensitivities in compound-specific strains. Although the sensitivity of lux strains can be enhanced by various molecular manipulations, most reported detection thresholds are still too high to detect levels of individual contaminants as they occur nowadays in European drinking waters. However, lux strains sensing specific toxic effects have the advantage of being able to respond to mixtures of contaminants inducing the same effect, and thus could be used as a sensor for the sum effect, including the effect of compounds that are as yet not identified by chemical analysis. An evaluation of the suitability of lux strains for monitoring surface and drinking water is therefore provided

Optimizing Effective Parameters to Enhance the Sensitivity of Vertical Flow Assay for Detection of Escherichia coli

Vertical flow immunoassays (VFIAs) are considered potential point-of-care testing (POCT) devices compared to lateral flow assays due to their ability to analyze a comparatively large sample volume and ease of multiplexing. However, VFIA devices are limited by low analytical sensitivity when coupled with a visual colorimetric signal. Herein, we carefully analyzed key parameters that accounted for the proper functionality of VFIA that can be modified to enhance the overall sensitivity of VFIA. In particular, we focused on improving the stability of conjugate pads impregnated with capture antibodies, maintaining a controlled flow rate to ensure higher analyte reactivity with capture antibodies, and enhancing the absorption efficiency. The results showed that air-drying of conjugate pads in the presence of 5% (w/v) lactose significantly improved the stability of antibodies during long-term storage. Integration of dissolvable polyvinyl alcohol (PVA) membrane of optimal concentration as a time-barrier film into the sensor delayed the flow of samples, thereby increasing the biorecognition interaction time between immunoreagents for the formation of immuno-complexes, which in turn led to higher sensitivity of the assay. Furthermore, the employment of an absorbent pad with higher water holding capacity significantly reduced the non-specific binding of immunocomplexes, thereby reducing the possibility of false-negative results

Smartphone-Based Whole-Cell Biosensor Platform Utilizing an Immobilization Approach on a Filter Membrane Disk for the Monitoring of Water Toxicants

Bioluminescent bacteria whole-cell biosensors (WCBs) have been widely used in a range of sensing applications in environmental monitoring and medical diagnostics. However, most of them use planktonic bacteria cells that require complicated signal measurement processes and therefore limit the portability of the biosensor device. In this study, a simple and low-cost immobilization method was examined. The bioluminescent bioreporter bacteria was absorbed on a filter membrane disk. Further optimization of the immobilization process was conducted by comparing different surface materials (polyester and parafilm) or by adding glucose and ampicillin. The filter membrane disks with immobilized bacteria cells were stored at −20 °C for three weeks without a compromise in the stability of its biosensing functionality for water toxicants monitoring. Also, the bacterial immobilized disks were integrated with smartphones-based signal detection. Then, they were exposed to water samples with ethanol, chloroform, and H2O2, as common toxicants. The sensitivity of the smartphone-based WCB for the detection of ethanol, chloroform, and H2O2 was 1% (v/v), 0.02% (v/v), and 0.0006% (v/v), respectively. To conclude, this bacterial immobilization approach demonstrated higher sensitivity, portability, and improved storability than the planktonic counterpart. The developed smartphone-based WCB establishes a model for future applications in the detection of environmental water toxicants

Biosensors based on combined optical and electrochemical transduction for molecular diagnostics

International audienc

Bioluminescence enhancement through an added washing protocol enabling a greater sensitivity to carbofuran toxicity

International audienceThe effects of carbofuran toxicity on a genetically modified bacterial strain E. coli DPD2794 were enhanced using a new bioluminescent protocol which consisted of three consecutive steps: incubation, washing and luminescence reading. Specifically, in the first step, several concentrations of carbofuran aqueous solutions were incubated with different bacterial suspensions at recorded optical densities for different lengths of time. Thereafter, the resulting bacterial/toxicant mixtures were centrifuged and the aged cellular supernatant replaced with fresh medium. In the final step, the carbofuran- induced bioluminescence to the exposed E. coli DPD2794 bacteria was shown to provide a faster and higher intensity when recorded at a higher temperature at30 °C which is not usually used in the literature. It was found that the incubation time and the replacement of aged cellular medium were essential factors to distinguish different concentrations of carbofuran in the bioluminescent assays. From our results, the optimum incubation time for a “light ON” bioluminescence detection of the effect of carbofuran was 6 h. Thanks to the replacement of the aged cellular medium, a group of additional peaks starting around 30 min were observed and we used the corresponding areas under the curve (AUC) at different contents of carbofuran to produce the calibration curve. Based on the new protocol, a carbofuran concentration of 0.5 pg/mL can be easily determined in a microtiter plate bioluminescent assay, while a non-wash protocol provides an unexplainable order of curve evolutionswhich does not allow the user to determine the concentration