Development of a flow-cell bioreactor for immobilized sulfidogenic sludge characterization using electrochemical H2S microsensors.

The sulfate-reduction process plays a crucial role in the biological valorization of SOx gases. However, a complete understanding of the sulfidogenic process in bioreactors is limited by the lack of technologies for characterizing the sulfate-reducing activity of immobilized biomass. In this work, we propose a flow-cell bioreactor (FCB) for characterizing sulfate-reducing biomass using H2S microsensors to monitor H2S production in real-time within a biofilm. To replace natural immobilization through extracellular polymeric substance production, sulfidogenic sludge was artificially immobilized using polymers. Physical and sulfate-reducing activity studies were performed to select a polymer-biomass matrix that maintained sulfate-reducing activity of biomass while providing strong microbial retention and mechanical strength. Several operational conditions of the sulfidogenic reactor allowed to obtain a H2S profiles under different inlet sulfate loads and, additionally, 3D mapping was assessed in order to perform a hydraulic characterization. Besides, the effects of artificial immobilization on biodiversity were investigated through the characterization of microbial communities. This study demonstrated the appropriateness of immobilized-biomass for characterization of sulfidogenic biomass in FCB using H2S electrochemical microsensors, and beneficial microbiological communities shifts as well as enrichment of sulfate-reducing bacteria have been confirmed.

Amperometric Inkjet-Printed Thyroxine Sensor Based on Customized Graphene and Tunned Cyclodextrins as the Preconcentration Element.

The determination of thyroid hormones has practical clinical significance for the diagnosis of hyperthyroidism and hypothyroidism diseases. Considering this aspect, a wide range of analytical methods for the detection of analytes, including immunoassay, chemiluminescence, mass spectroscopy and high-performance liquid chromatography, among others, has been developed. This type of analysis provides feasible results. Nevertheless, it requires qualified staff, special facilities and is time-consuming. For this reason, this paper relies on the fabrication of an electrochemical device developed with inkjet printing technology for the free detection of Thyroxine (T4). To manufacture our electrochemical device, several aspects were considered from the use of materials that amplify electrical signals, to finding a supramolecular scaffold that possess affinity towards the target analyte and the need of preconcentrating the analyte on the electrode's surface. For this task, printed devices were modified with a hybrid nanomaterial consisting of reduced graphene oxide (rGO) tuned with Au nanoparticles (Au-NPs) and an entrapment agent and different thiolated cyclodextrins (x-CD-SH) as carrying agents. Analytes were preconcentrated via supramolecular chemistry due to the formation of an inclusion complex between the cyclodextrin and hormones. Morphological and electrochemical characterization of the final device was carried out to ensure the proper workability of the electrode, achieving excellent response, sensitivity and limit of detection (LOD).

Feasible H2S Sensing in Water with a Printed Amperometric Microsensor.

Concern over pollution has led to an increase in wastewater treatment systems, which require constant monitorization. In particular, hydrogen sulfide (H2S) is a toxic gas, soluble in water, commonly found in industrial and urban effluents. For proper removal control, fast, durable, and easy-to-handle analytical systems, capable of on-line measurements, such as electrochemical sensors, are required. Moreover, for a proper monitoring of said treatment processes, analysis must be carried out through all steps, thus needing for an economic and highly reproducible method of sensor fabrication. Digital printing have risen in the last few years as technologies capable of mass producing miniaturized electronical devices, allowing for the fabrication of amperometric sensors. Here, a 2 mm2 graphite (Gr) electrode, modified with different dispersions of single-walled carbon nanotubes (SWCNTs), poly(vinyl alcohol), poly(diallyl dimethylammonium chloride), and polylactic acid (PLA), is presented as a H2S sensor. SWCNTs allow for lower oxidation potentials, higher sensitivity, and a reduced rate of sulfur poisoning, while polymer dispersion of PLA increases mechanical stability and as a result, electrochemical performance. This microsensor presents an optimal pH working range between 7.5 and 11.0, a limit of detection of 4.3 µM, and the capacity to operate on complex matrices for H2S contamination detection.

Fully Inkjet-Printed Biosensors Fabricated with a Highly Stable Ink Based on Carbon Nanotubes and Enzyme-Functionalized Nanoparticles.

Enzyme inks can be inkjet printed to fabricate enzymatic biosensors. However, inks containing enzymes present a low shelf life because enzymes in suspension rapidly lose their catalytic activity. Other major problems of printing these inks are the non-specific adsorption of enzymes onto the chamber walls and stability loss during printing as a result of thermal and/or mechanical stress. It is well known that the catalytic activity can be preserved for significantly longer periods of time and to harsher operational conditions when enzymes are immobilized onto adequate surfaces. Therefore, in this work, horseradish peroxidase was covalently immobilized onto silica nanoparticles. Then, the nanoparticles were mixed into an aqueous ink containing single walled carbon nanotubes. Electrodes printed with this specially formulated ink were characterized, and enzyme electrodes were printed. To test the performance of the enzyme electrodes, a complete amperometric hydrogen peroxide biosensor was fabricated by inkjet printing. The electrochemical response of the printed electrodes was evaluated by cyclic voltammetry in solutions containing redox species, such as hexacyanoferrate (III/II) ions or hydroquinone. The response of the enzyme electrodes was studied for the amperometric determination of hydrogen peroxide. Three months after the ink preparation, the printed enzyme electrodes were found to still exhibit similar sensitivity, demonstrating that catalytic activity is preserved in the proposed ink. Thus, enzyme electrodes can be successfully printed employing highly stable formulation using nanoparticles as carriers.

Specially Designed Polyaniline/Polypyrrole Ink for a Fully Printed Highly Sensitive pH Microsensor.

pH sensing for healthcare applications requires sensors with mechanically stable materials of high sensitivity and high reproducibility combined with low-cost fabrication technologies. This work proposes a fully printed pH sensor based on a specially formulated conducting polymer deposited on a microelectrode in a flexible substrate. A formulation, which combined polyaniline (PANI) and polypyrrole (PPy) with integrated polyelectrolyte poly(sodium 4-styrenesulfonate) (PSS), was specially prepared to be printed by inkjet printing (IJP). The sensor has good sensitivity in the physiological region (pH 7-7.5) key for the healthcare biosensor. This mixture printed over a commercial gold ink, which has a singular chemical functionalization with phthalocyanine (Pc), increased the sensor sensitivity, showing an excellent reproducibility with a linear super-Nernstian response (81.2 ± 0.5 mV/pH unit) in a wide pH range (pH 3-10). This new ink together with the IJP low-cost technique opens new opportunities for pH sensing in the healthcare field with a single device, which is disposable, highly sensitive, and stable in the whole pH range.

Gut-on-a-chip: Mimicking and monitoring the human intestine.

Over the last few years, the intestine has been extensively studied using in vitro microfluidic systems, commonly known as gut-on-a-chip (GOC) devices. This interest has been due not only to the importance of the intestine's proper functions but also to the relationship that this organ and the microbiota that inhabits it has with the rest of the body's organs. The increased complexity of these in vitro systems, together with the need to improve our understanding of intestinal physiology interdependencies, has led to greater focus on the integration of biosensors within these devices. However, the current number of GOC devices with integrated sensors for monitoring relevant physiological parameters are very limited and demand the use of external analytical techniques that delay the analysis and prevent real-time decision-making. This paper reviews the various materials, technologies, and structures that have been used both for mimicking the physiology of the intestine and monitoring relevant physiological parameters, such as permeability of the gut barrier, dissolved oxygen concentration, cytokines profile and the production of microbial short-chain fatty acids. We also propose alternative biosensing techniques demonstrated in other in vitro and lab-on-a-chip devices that could be translated to GOC models. A critical analysis of the requirements, limitations, and current challenges on the microenvironment replication and monitorization of GOC models is included, with a particular focus on the physiological parameters and biomarkers that should be detected simultaneously in real-time to get a proper framework of the gut function that until now, have not received the necessary attention.

Cortical Superficial Siderosis: A Descriptive Analysis in a Memory Clinic Population.

BACKGROUND: Cortical superficial siderosis (cSS) is a hemorrhagic marker of blood-brain barrier disruption detected in brain MRI. Together with cerebral microbleeds (CMB), they are recognized as a small vessel disease marker associated with cerebral amyloid angiopathy. OBJECTIVE: This study aims to determine the prevalence and the characteristics of cSS in a memory clinic population. METHODS: Cross-sectional retrospective analysis of 613 patients from Geneva University Hospitals memory clinic. All patients underwent standardized brain MRI and neuropsychological assessment with diagnosis confirmed by an expert. The presence of cSS was visually assessed and classified as focal (restricted to 3 sulci) or disseminated within the correspondent topography. CMB were classified according to the Microbleed Anatomical Rating Scale. RESULTS: cSS was detected in 26/613 patients (4.2%), classified as disseminated in 5/26 cases (19%). Alzheimer's disease (AD) and AD associated with a significant vascular component were the diagnoses more frequently related to cSS (18/26; 69%). Patients with cSS had an increased prevalence of both hypertension (81% versus 57%; p = 0.015) and WMH burden (p = 0.012). The overall prevalence of cerebral microbleeds (69% versus 32%; p < 0.01), as well as their mean number (0.69±0.47 versus 0.32±0.46; p < 0.01) were both increased in patients with cSS. In the logistic regression model, the presence of 5 or more CMB (OR 11.35; 95% CI 4.68-27.55; p < 0.01) and hypertension (OR 3.31; 95% CI 1.19-9.15; p = 0.021) were significantly associated with cSS. CONCLUSIONS: cSS is observed in patients diagnosed with AD and AD with a vascular component, being independently associated with multiple CMB and hypertension.

A Minimally Invasive Microsensor Specially Designed for Simultaneous Dissolved Oxygen and pH Biofilm Profiling.

A novel sensing device for simultaneous dissolved oxygen (DO) and pH monitoring specially designed for biofilm profiling is presented in this work. This device enabled the recording of instantaneous DO and pH dynamic profiles within biofilms, improving the tools available for the study and the characterization of biological systems. The microsensor consisted of two parallel arrays of microelectrodes. Microelectrodes used for DO sensing were bare gold electrodes, while microelectrodes used for pH sensing were platinum-based electrodes modified using electrodeposited iridium oxide. The device was fabricated with a polyimide (Kapton®) film of 127 µm as a substrate for minimizing the damage caused on the biofilm structure during its insertion. The electrodes were covered with a Nafion® layer to increase sensor stability and repeatability and to avoid electrode surface fouling. DO microelectrodes showed a linear response in the range 0-8 mg L-1, a detection limit of 0.05 mg L-1, and a sensitivity of 2.06 nA L mg-1. pH electrodes showed a linear super-Nernstian response (74.2 ± 0.7 mV/pH unit) in a wide pH range (pH 4-9). The multi-analyte sensor array was validated in a flat plate bioreactor where simultaneous and instantaneous pH and DO profiles within a sulfide oxidizing biofilm were recorded. The electrodes spatial resolution, the monitoring sensitivity, and the minimally invasive features exhibited by the proposed microsensor improved biofilm monitoring performance, enabling the quantification of mass transfer resistances and the assessment of biological activity.

Stable Full-Inkjet-Printed Solid-State Ag/AgCl Reference Electrode.

With a growing demand for the availability of inexpensive, simple, and rapid prototyped devices, the prospect of miniaturization of the reference electrodes using printing techniques becomes promising. A stable and reusable full-inkjet-printed solid-state reference electrode (IPRE) was developed. The reference electrode was fully produced by consecutive inkjet printing of several layers. Ag ink was printed and chlorinated by NaClO printing, forming a Ag/AgCl pseudoreference electrode. Then a surface protection by printing a Cl--saturated polyvinyl butyral membrane finally gave a reference electrode that demonstrated an outstanding performance comparable to commercial ones. This full inkjet printing fabrication strategy will improve the viability of producing low-cost miniaturized reference electrodes with interest in many electrochemical sensor-dependent areas.

Enhanced Performance Stability of Iridium Oxide-Based pH Sensors Fabricated on Rough Inkjet-Printed Platinum.

Today, electrochemical sensors are used for a broad range of applications. A fundamental challenge is still the achievement of long-term sensor stability by ensuring good adhesion between the deposited sensing layer and the substrate material, e.g., a metal electrode. Until now, the most applied strategy to overcome this problem is to increase the surface roughness of the metal layer by mechanical etching or by electroplating of additional material layers, which both imply an increase in manufacturing steps and thus the final cost of the overall device. Alternatively, to overcome these adhesion problems, we propose the direct printing of a novel platinum nanoparticle ink, which is compatible with low-cost additive digital inkjet and with flexible low-cost substrates. This water-based platinum ink has two unique features: it leads to highly rough surfaces, which promotes the adhesion of deposited sensing material, and it is a highly low-temperature curing ink, compatible with polymeric substrates that cannot withstand high temperatures. Based on this concept, we report about a long-term stable and highly sensitive solid-state pH sensor functionalized by anodic electrodeposited iridium oxide on a rough nanostructured platinum printed layer. The sensors showed an excellent reproducibility with a linear super-Nernstian response (71.3 ± 0.3 mV/pH unit) in a wide pH range (pH 2-11). Long-term stability tests for over 1 year of application demonstrate an excellent mechanical sensor layer stability, which is correlated to the distinct roughness of the printed platinum layer. This novel approach is useful to simplify the fabrication process and with that the sensor costs.

Quantification of Signal-to-Noise Ratio in Cerebral Cortex Recordings Using Flexible MEAs With Co-localized Platinum Black, Carbon Nanotubes, and Gold Electrodes.

Developing new standardized tools to characterize brain recording devices is critical to evaluate neural probes and for translation to clinical use. The signal-to-noise ratio (SNR) measurement is the gold standard for quantifying the performance of brain recording devices. Given the drawbacks with the SNR measure, our first objective was to devise a new method to calculate the SNR of neural signals to distinguish signal from noise. Our second objective was to apply this new SNR method to evaluate electrodes of three different materials (platinum black, Pt; carbon nanotubes, CNTs; and gold, Au) co-localized in tritrodes to record from the same cortical area using specifically designed multielectrode arrays. Hence, we devised an approach to calculate SNR at different frequencies based on the features of cortical slow oscillations (SO). Since SO consist in the alternation of silent periods (Down states) and active periods (Up states) of neuronal activity, we used these as noise and signal, respectively. The spectral SNR was computed as the power spectral density (PSD) of Up states (signal) divided by the PSD of Down states (noise). We found that Pt and CNTs electrodes have better recording performance than Au electrodes for the explored frequency range (5-1500 Hz). Together with two proposed SNR estimators for the lower and upper frequency limits, these results substantiate our SNR calculation at different frequency bands. Our results provide a new validated SNR measure that provides rich information of the performance of recording devices at different brain activity frequency bands (<1500 Hz).

Inkjet-Printed Sulfide-Selective Electrode.

Inkjet printing technology has emerged as an alternative manufacturing method for low-cost production of electrodes. Despite significant progress, there is still a lack in the production of ion-selective electrodes. Herein, the two-step fabrication of the first inkjet-printed sulfide-selective electrode (IPSSE) is described. The two-step fabrication consists of printing a silver electrode followed by an electrochemical deposition of sulfide to produce a second kind electrode (Ag/Ag2S). The performance of this novel device was tested using potentiometric measurements. Nernstian response (-29.4 ± 0.3 mV·decade-1) was obtained within concentrations of 0.03-50 mM with a response time of ∼3 s. Furthermore, river/sea-spiked environmental samples and samples from a bioreactor for sulfate reduction to sulfide were measured and compared against a commercial sensor giving no significant differences. The IPSSE described in this work showed good reproducibility and durability during daily measurements over 15 days without any special storage conditions. Considering all the current challenges in inkjet-printed ion-selective electrodes, this different fabrication approach opens a new perspective for mass production of all-solid state ion-selective electrodes.

Dynamic characterization of external and internal mass transport in heterotrophic biofilms from microsensors measurements.

Knowledge of mass transport mechanisms in biofilm-based technologies such as biofilters is essential to improve bioreactors performance by preventing mass transport limitation. External and internal mass transport in biofilms was characterized in heterotrophic biofilms grown on a flat plate bioreactor. Mass transport resistance through the liquid-biofilm interphase and diffusion within biofilms were quantified by in situ measurements using microsensors with a high spatial resolution (<50 µm). Experimental conditions were selected using a mathematical procedure based on the Fisher Information Matrix to increase the reliability of experimental data and minimize confidence intervals of estimated mass transport coefficients. The sensitivity of external and internal mass transport resistances to flow conditions within the range of typical fluid velocities over biofilms (Reynolds numbers between 0.5 and 7) was assessed. Estimated external mass transfer coefficients at different liquid phase flow velocities showed discrepancies with studies considering laminar conditions in the diffusive boundary layer near the liquid-biofilm interphase. The correlation of effective diffusivity with flow velocities showed that the heterogeneous structure of biofilms defines the transport mechanisms inside biofilms. Internal mass transport was driven by diffusion through cell clusters and aggregates at Re below 2.8. Conversely, mass transport was driven by advection within pores, voids and water channels at Re above 5.6. Between both flow velocities, mass transport occurred by a combination of advection and diffusion. Effective diffusivities estimated at different biofilm densities showed a linear increase of mass transport resistance due to a porosity decrease up to biofilm densities of 50 g VSS·L(-1). Mass transport was strongly limited at higher biofilm densities. Internal mass transport results were used to propose an empirical correlation to assess the effective diffusivity within biofilms considering the influence of hydrodynamics and biofilm density.

Biofilm dynamics characterization using a novel DO-MEA sensor: mass transport and biokinetics.

Biodegradation process modeling is an essential tool for the optimization of biotechnologies related to gaseous pollutant treatment. In these technologies, the predominant role of biofilm, particularly under conditions of no mass transfer limitations, results in a need to determine what processes are occurring within the same. By measuring the interior of the biofilms, an increased knowledge of mass transport and biodegradation processes may be attained. This information is useful in order to develop more reliable models that take biofilm heterogeneity into account. In this study, a new methodology, based on a novel dissolved oxygen (DO) and mass transport microelectronic array (MEA) sensor, is presented in order to characterize a biofilm. Utilizing the MEA sensor, designed to obtain DO and diffusivity profiles with a single measurement, it was possible to obtain distributions of oxygen diffusivity and biokinetic parameters along a biofilm grown in a flat plate bioreactor (FPB). The results obtained for oxygen diffusivity, estimated from oxygenation profiles and direct measurements, revealed that changes in its distribution were reduced when increasing the liquid flow rate. It was also possible to observe the effect of biofilm heterogeneity through biokinetic parameters, estimated using the DO profiles. Biokinetic parameters, including maximum specific growth rate, the Monod half-saturation coefficient of oxygen, and the maintenance coefficient for oxygen which showed a marked variation across the biofilm, suggest that a tool that considers the heterogeneity of biofilms is essential for the optimization of biotechnologies.

SU-8 based microprobes for simultaneous neural depth recording and drug delivery in the brain.

While novel influential concepts in neuroscience bring the focus to local activities generated within a few tens of cubic micrometers in the brain, we are still devoid of appropriate tools to record and manipulate pharmacologically neuronal activity at this fine scale. Here we designed, fabricated and encapsulated microprobes for simultaneous depth recording and drug delivery using exclusively the polymer SU-8 as structural material. A tetrode- and linear-like electrode patterning was combined for the first time with single and double fluidic microchannels for independent drug delivery. The device was tested experimentally using the in vivo anesthetized rat preparation. Both probe types successfully recorded detailed spatiotemporal features of local field potentials and single-cell activity at a resolution never attained before with integrated fluidic probes. Drug delivery was achieved with high spatial and temporal precision in a range from tens of nanoliters to a few microliters, as confirmed histologically. These technological advancements will foster a wide range of neural applications aimed at simultaneous monitoring of brain activity and delivery at a very precise micrometer scale.

Vertically aligned carbon nanotubes for microelectrode arrays applications.

In this work a methodology to fabricate carbon nanotube based electrodes using plasma enhanced chemical vapour deposition has been explored and defined. The final integrated microelectrode based devices should present specific properties that make them suitable for microelectrode arrays applications. The methodology studied has been focused on the preparation of highly regular and dense vertically aligned carbon nanotube (VACNT) mat compatible with the standard lithography used for microelectrode arrays technology.

SU-8 based microprobes with integrated planar electrodes for enhanced neural depth recording.

Here, we describe new fabrication methods aimed to integrate planar tetrode-like electrodes into a polymer SU-8 based microprobe for neuronal recording applications. New concepts on the fabrication sequences are introduced in order to eliminate the typical electrode-tissue gap associated to the passivation layer. Optimization of the photolithography technique and high step coverage of the sputtering process have been critical steps in this new fabrication process. Impedance characterization confirmed the viability of the electrodes for reliable neuronal recordings with values comparable to commercial probes. Furthermore, a homogeneous sensing behavior was obtained in all the electrodes of each probe. Finally, in vivo action potential and local field potential recordings were successfully obtained from the rat dorsal hippocampus. Peak-to-peak amplitude of action potentials ranged from noise level to up to 400-500 µV. Moreover, action potentials of different amplitudes and shapes were recorded from all the four recording sites, suggesting improved capability of the tetrode to distinguish from different neuronal sources.

Cancer prognostics by direct detection of p53-antibodies on gold surfaces by impedance measurements.

The identification and measurement of biomarkers is critical to a broad range of methods that diagnose and monitor many diseases. Serum auto-antibodies are rapidly becoming interesting targets because of their biological and medical relevance. This paper describes a highly sensitive, label-free approach for the detection of p53-antibodies, a prognostic indicator in ovarian cancer as well as a biomarker in the early stages of other cancers. This approach uses impedance measurements on gold microelectrodes to measure antibody concentrations at the picomolar level in undiluted serum samples. The biosensor shows high selectivity as a result of the optimization of the epitopes responsible for the detection of p53-antibodies and was validated by several techniques including microcontact printing, self-assembled-monolayer desorption ionization (SAMDI) mass spectrometry, and adhesion pull-off force by atomic force microscopy (AFM). This transduction method will lead to fast and accurate diagnostic tools for the early detection of cancer and other diseases.

Magnetic entrapment for fast, simple and reversible electrode modification with carbon nanotubes: application to dopamine detection.

Carbon nanotubes (CNT) have been exploited for an important number of electroanalytical and sensing purposes. Specifically, CNT incorporation to an electrode surface coating increases its roughness and area, provides electrocatalytic activity towards a variety of molecules, and improves electron transfer. This modification is generally based on the irreversible deposition of CNT on surface. Nevertheless, CNT are highly porous materials that might promote molecule non-specific adsorption and/or electrodeposition, which could induce sample-to-sample cross-contamination and affect measurement specificity and reproducibility. This drawback has been often circumvented by combining CNT with charged polymers able to repel molecules of opposed charge. We demonstrate that single-walled CNT (SWCNT) have a strong tendency to non-specifically adsorb onto the surface of protein-coated magnetic particles (MP). Magnetic capture of those MP generates CNT coentrapment and allows extremely fast, simple and reversible production of SWCNT electrodes. We have exploited this phenomenon for the production of modified screen-printed electrodes (MP/CNT-SPE), which have been characterized by Scanning Electron Microscopy. The surface has been additionally optimized by evaluating the electrochemical performance of SPE modified with different amounts and proportions of MP and CNT. The modified devices have then been used for dopamine detection. MP/CNT-SPE generated improved assay sensitivity, lower limit of detection, and up to 500% higher current signals than bare electrodes. Magnetic entrapment is proposed as a promising strategy for the fast, simple and reversible generation of nanostructured electrodes of enhanced performance within a few minutes and electrode re-utilisation by simple magnet removal and surface washing.

Easily made single-walled carbon nanotube surface microelectrodes for neuronal applications.

The present work examines the feasibility of a simple method for using single-walled carbon nanotubes (SWNT) to fabricate multielectrode arrays (MEA) for electrophysiological recordings. A suspension of purified SWNTs produced by arc discharged was directly deposited onto standard platinum electrodes. The in vitro impedance and electrochemical characterizations demonstrated the enhanced electrical properties of the SWNT microelectrode array. To test its functionality we performed extracellular ganglion cell recordings in isolated superfused rabbit retinas. Our results showed that SWNT based electrode arrays have potential advantages over metal electrodes and can be successfully used to record the single and multi-unit activity of ganglion cell populations.