In Situ Performance Monitoring of Electrochemical Oxygen and Hydrogen Peroxide Sensors in an Additively Manufactured Modular Microreactor.

Miniaturized and microstructured reactors in process engineering are essential for a more decentralized, flexible, sustainable, and resilient chemical production. Modern, additive manufacturing methods for metals enable complex reactor-geometries, increased functionality, and faster design iterations, a clear advantage over classical subtractive machining and polymer-based approaches. Integrated microsensors allow online, in situ process monitoring to optimize processes like the direct synthesis of hydrogen peroxide. We developed a modular tube-in-tube membrane reactor fabricated from stainless steel via 3D printing by laser powder bed fusion of metals (PBF-LB/M). The reactor concept enables the spatially separated dosage and resaturation of two gaseous reactants across a membrane into a liquid process medium. Uniquely, we integrated platinum-based electrochemical sensors for the online detection of analytes to reveal the dynamics inside the reactor. An advanced chronoamperometric protocol combined the simultaneous concentration measurement of hydrogen peroxide and oxygen with monitoring of the sensor performance and self-calibration in long-term use. We demonstrated the highly linear and sensitive monitoring of hydrogen peroxide and dissolved oxygen entering the liquid phase through the membrane. Our measurements delivered important real-time insights into the dynamics of the concentrations in the reactor, highlighting the power of electrochemical sensors applied in process engineering. We demonstrated the stable continuous measurement over 1 week and estimated the sensor lifetime for months in the acidic process medium. Our approach combines electrochemical sensors for process monitoring with advanced, additively manufactured stainless steel membrane microreactors, supporting the power of sensor-equipped microreactors as contributors to the paradigm change in process engineering and toward a greener chemistry.

Electrochemical Methods in the Cloud: FreiStat, an IoT-Enabled Embedded Potentiostat.

Embedded potentiostats enable electrochemical measurements in the Internet-of-Things (IoT) or other decentralized applications, such as remote environmental monitoring, electrochemical energy systems, and biomedical point-of-care applications. We report on Freiburg's Potentiostat (FreiStat) based on the AD5941 potentiostat circuit from Analog Devices, together with custom firmware, as the key to precise and advanced electrochemical methods. We demonstrated its analytical performance by various cyclic voltammetry measurements, advanced techniques such as differential pulse voltammetry, and a lactate biosensor measurement with currents in the nA range and a resolution of 54 pA. The FreiStat yielded analytical results comparable to benchtop devices and outperformed current commercial embedded potentiostats at significantly lower cost, smaller size, and lower power consumption. Decentralized corrosion analysis by a Tafel plot using the IBM Cloud showed its applicability in a typical IoT scenario. The developed open-source software framework facilitates the integration of electrochemical instrumentation into applications utilizing machine learning and other artificial intelligence. Together with the affordable and highly capable embedded potentiostat, our approach can leverage analytical chemistry toward increasingly important, more widespread and decentralized applications.

Advanced electrochemical potential monitoring for improved understanding of electrical neurostimulation protocols.

Objective.Current-controlled neurostimulation is increasingly used in the clinical treatment of neurological disorders and widely applied in neural prostheses such as cochlear implants. Despite its importance, time-dependent potential traces of electrodes during microsecond-scale current pulses, especially with respect to a reference electrode (RE), are not precisely understood. However, this knowledge is critical to predict contributions of chemical reactions at the electrodes, and ultimately electrode stability, biocompatibility, and stimulation safety and efficacy.Approach.We assessed the electrochemistry of neurostimulation protocolsin vitrowith Pt microelectrodes from millisecond (classical electroanalysis) to microsecond (neurostimulation) timescales. We developed a dual-channel instrumentation amplifier to include a RE in neurostimulation setups. Uniquely, we combined potential measurements with potentiostatic prepolarization to control and investigate the surface status, which is not possible in typical stimulation setups.Main results.We thoroughly validated the instrumentation and highlighted the importance of monitoring individual electrochemical electrode potentials in different configurations of neurostimulation. We investigated electrode processes such as oxide formation and oxygen reduction by chronopotentiometry, bridging the gap between milli- and microsecond timescales. Our results demonstrate how much impact on potential traces the electrode's initial surface state and electrochemical surface processes have, even on a microsecond scale.Significance.Our unique use of preconditioning in combination with stimulation reveals that interpreting potential traces with respect to electrode processes is misleading without rigorous control of the electrode's surface state. Especiallyin vivo, where the microenvironment is unknown, simply measuring the voltage between two electrodes cannot accurately reflect the electrode's state and processes. Potential boundaries determine charge transfer, corrosion, and alterations of the electrode/tissue interface such as pH and oxygenation, particularly in long-termin vivouse. Our findings are relevant for all use-cases of constant-current stimulation, strongly advocating for electrochemicalin situinvestigations in many applications like the development of new electrode materials and stimulation methods.

Multiplexed biosensor for point-of-care COVID-19 monitoring: CRISPR-powered unamplified RNA diagnostics and protein-based therapeutic drug management.

In late 2019 SARS-CoV-2 rapidly spread to become a global pandemic, therefore, measures to attenuate chains of infection, such as high-throughput screenings and isolation of carriers were taken. Prerequisite for a reasonable and democratic implementation of such measures, however, is the availability of sufficient testing opportunities (beyond reverse transcription PCR, the current gold standard). We, therefore, propose an electrochemical, microfluidic multiplexed polymer-based biosensor in combination with CRISPR/Cas-powered assays for low-cost and accessible point-of-care nucleic acid testing. In this study, we simultaneously screen for and identify SARS-CoV-2 infections (Omicron-variant) in clinical specimens (Sample-to-result time: ∼30 min), employing LbuCas13a, whilst bypassing reverse transcription as well as target amplification of the viral RNA (LODs of 2,000 and 7,520 copies/µl for the E and RdRP genes, respectively, and 50 copies/ml for combined targets), both of which are necessary for detection via PCR and other isothermal methods. In addition, we demonstrate the feasibility of combining synthetic biology-driven assays based on different classes of biomolecules, in this case protein-based ß-lactam antibiotic detection, on the same device. The programmability of the effector and multiplexing capacity (up to six analytes) of our platform, in combination with a miniaturized measurement setup, including a credit card sized near field communication (NFC) potentiostat and a microperistaltic pump, provide a promising on-site tool for identifying individuals infected with variants of concern and monitoring their disease progression alongside other potential biomarkers or medication clearance.

Bioprinting-based automated deposition of single cancer cell spheroids into oxygen sensor microelectrode wells.

Three-dimensional (3D) cell agglomerates, such as microtissues, organoids, and spheroids, become increasingly relevant in biomedicine. They can provide in vitro models that recapitulate functions of the original tissue in the body and have applications in cancer research. For example, they are widely used in organ-on-chip systems. Microsensors can provide essential real-time information on cell metabolism as well as the reliability and quality of culture conditions. The combination of sensors and 3D cell cultures, especially single spheroids, is challenging in terms of reproducible formation, manipulation, and access to spheroids, precise positioning near sensors, and high cell-to-volume ratios to obtain meaningful biosignals in the most parallel approach possible. To overcome this challenge, we combined state-of-the-art bioprinting techniques to automatically print tumour spheroids directly into microwells of a chip-based electrochemical oxygen sensor array. We demonstrated highly accurate and reproducible spheroid formation (diameter of approx. 200 µm) and a spheroid deposition precision of 25 µm within a volume of 22 nl per droplet. Microstructures and hydrogel-coated microwells allowed the placement of single MCF-7 breast cancer spheroids close to the sensor electrodes. The microelectrode wells were sealed for oxygen measurements within a 55 nl volume for fast concentration changes. Accurate and stable amperometric oxygen sensor performance was demonstrated from atmospheric to anoxic regions. Cellular respiration rates from single tumour spheroids in the range of 450-850 fmol min-1 were determined, and alterations of cell metabolism upon drug exposure were shown. Our results uniquely combine bioprinting with 3D cell culture monitoring and demonstrate the much-needed effort for facilitation, parallelization, sensor integration, and drug delivery in 3D cell culture and organ-on-chip experiments. The workflow has a high degree of automation and potential for scalability. In order to achieve greater flexibility in the automation of spheroid formation and trapping, we employed a method based on drop-on-demand liquid handling systems, instead of the typical on-chip approach commonly used in microfluidics. Its relevance ranges from fundamental metabolic research over standardization of cell culture experiments and toxicological studies, to personalized medicine, e.g. patient-specific chemotherapy.

In situ stability monitoring of platinum thin-film electrodes for neural interfaces in the presence of proteins.

The long-term stability of platinum electrodes is a key factor that determines the life-time of biomedical devices, such as implanted neural interfaces like brain stimulation or recording electrodes, cochlear implants, and biosensors. The downsizing of such devices relies on the usage of microfabricated thin-film electrodes. In order to determine and investigate the causal degradation processes for platinum electrodes, it is essential to use potential-controlled experiments, which allow selectable polarization of the electrode without exceeding the water stability window boundaries. Therefore, the surface processes and redox reactions occurring at the electrode are known at all times. In this study, we present the continuous in situ monitoring of platinum-based thin-film electrodes along their complete life cycle in neutral pH with and without the presence of proteins. The usage of chronoamperometry for electrode aging, monitoring of surface processes and the tracking of analyte redox processes, together with cyclic voltammetry to determine the complete amount of surface charge, allows a reliable quantification of fundamental degradation processes. We found that platinum dissolution is primarily driven by the formation and removal of Pt oxide. Despite the significantly lowered charge transfer, the presence of proteins did not prevent material loss or increase electrode lifetime. These results should be considered when interpreting results from current-controlled methods as typically used for neural interfaces. Clinical Relevance- All clinically relevant applications of microelectrodes, ranging from cell culture over diagnostics to in vivo use, involve the presence of proteins. Detailed and fundamental insight into electrode stability in the presence of proteins is therefore essential for successful clinical translation of neural interface technologies.

Electrochemical microelectrode degradation monitoring:in situinvestigation of platinum corrosion at neutral pH.

Objective. The stability of platinum and other noble metal electrodes is critical for neural implants, electrochemical sensors, and energy sources. Beyond the acidic or alkaline environment found in most electrochemical studies, the investigation of electrode corrosion in neutral pH and chloride containing electrolytes is essential, particularly regarding the long-term stability of neural interfaces, such as brain stimulation electrodes or cochlear implants. In addition, the increased use of microfabricated devices demands the investigation of thin-film electrode stability in combination with electrode performance.Approach. We developed a procedure of electrochemical methods for continuous tracking of electrode degradationin situover the complete life cycle of platinum thin-film microelectrodes in a unique combination with simultaneous chemical sensing. We used chronoamperometry and cyclic voltammetry to measure electrode surface and analyte redox processes, together with accelerated electrochemical degradation.Main results.We compared degradation between thin-film microelectrodes and bulk electrodes, neutral to acidic pH, different pulsing schemes, and the presence of the redox active species oxygen and hydrogen peroxide. Results were confirmed by electrochemical impedance spectroscopy, as well as mechanical profilometry and microscopy to determine material changes on a nanometer scale. We found that electrode degradation is mainly driven by repeated formation and removal of the platinum surface oxide, also within the electrochemical stability window of water. There was no considerable difference between thin-film micro- and macroscopic bulk electrodes or in the presence of reactive species, whereas acidic pH or extending the potential window led to increased degradation.Significance.Our results provide valuable fundamental information on platinum microelectrode degradation under conditions found in biomedical applications. For the first time, we employed a unified method to report quantitative data on electrode degradation up to a defined endpoint. Our method is a widely applicable framework for comparative long-term studies of electrode micro-/nanomaterial, sensor and neural interface stability.

Biosensor-Enabled Multiplexed On-Site Therapeutic Drug Monitoring of Antibiotics.

Personalized antibiotherapy ensures that the antibiotic concentration remains in the optimal therapeutic window to maximize efficacy, minimize side effects, and avoid the emergence of drug resistance due to insufficient dosing. However, such individualized schemes need frequent sampling to tailor the blood antibiotic concentrations. To optimally integrate therapeutic drug monitoring (TDM) into the clinical workflow, antibiotic levels can either be measured in blood using point-of-care testing (POCT), or can rely on noninvasive sampling. Here, a versatile biosensor with an antibody-free assay for on-site TDM is presented. The platform is evaluated with an animal study, where antibiotic concentrations are quantified in different matrices including whole blood, plasma, urine, saliva, and exhaled breath condensate (EBC). The clearance and the temporal evaluation of antibiotic levels in EBC and plasma are demonstrated. Influence of matrix effects on measured drug concentrations is determined by comparing the plasma levels with those in noninvasive samples. The system's potential for blood-based POCT is further illustrated by tracking ß-lactam concentrations in untreated blood samples. Finally, multiplexing capabilities are explored successfully for multianalyte/sample analysis. By enabling a rapid, low-cost, sample-independent, and multiplexed on-site TDM, this system can shift the paradigm of "one-size-fits-all" strategy.

Standard cochlear implants as electrochemical sensors: Intracochlear oxygen measurements in vivo.

Cochlear implants are the most successful neural prostheses worldwide and routinely restore sensorineural hearing loss by direct electrical stimulation of the auditory nerve. Enhancing this standard implant by chemical sensor functionality opens up new possibilities, ranging from access to the biochemical microenvironment of the implanted electrode array to the long-term study of the electrode status. We developed an electrochemical method to turn the platinum stimulation microelectrodes of cochlear implants into electrochemical sensors. The electrodes showed excellent and stable chemical sensor properties, as demonstrated by in vitro characterizations with combined amperometric and active potentiometric dissolved oxygen and hydrogen peroxide measurements. Linear, stable and highly reproducible sensor responses within the relevant concentration ranges with negligible offset were shown. This approach was successfully applied in vivo in an animal model. Intracochlear oxygen dynamics in rats upon breathing pure oxygen were reproducibly and precisely measured in real-time from the perilymph. At the same time, correct implant placement and its functionality was verified by measurements of electrically evoked auditory brainstem responses with clearly distinguishable peaks. Acute measurements indicated no adverse influence of electrical stimulation on electrochemical measurements and vice versa. Our work is ground-breaking towards advanced implant functionality, future implant lifetime monitoring, and implant-life-long in situ investigation of electrode degradation in cochlear implant patients.

Microfluidic organ-on-chip system for multi-analyte monitoring of metabolites in 3D cell cultures.

Three-dimensional cell cultures using patient-derived stem cells are essential in vitro models for a more efficient and individualized cancer therapy. Currently, culture conditions and metabolite concentrations, especially hypoxia, are often not accessible continuously and in situ within microphysiological systems. However, understanding and standardizing the cellular microenvironment are the key to successful in vitro models. We developed a microfluidic organ-on-chip platform for matrix-based, heterogeneous 3D cultures with fully integrated electrochemical chemo- and biosensor arrays for the energy metabolites oxygen, lactate, and glucose. Advanced microstructures allow straightforward cell matrix integration with standard laboratory equipment, compartmentalization, and microfluidic access. Single, patient-derived, triple-negative breast cancer stem cells develop into tumour organoids in a heterogeneous spheroid culture on-chip. Our system allows unprecedented control of culture conditions, including hypoxia, and simultaneous verification by integrated sensors. Beyond previous works, our results demonstrate precise and reproducible on-chip multi-analyte metabolite monitoring under dynamic conditions from a matrix-based culture over more than one week. Responses to alterations in culture conditions and cancer drug exposure, such as metabolite consumption and production rates, could be accessed quantitatively and in real-time, in contrast to endpoint analyses. Our approach highlights the importance of continuous, in situ metabolite monitoring in 3D cell cultures regarding the standardization and control of culture conditions, and drug screening in cancer research. Overall, the results underline the potential of microsensors in organ-on-chip systems for successful application, e.g. in personalized medicine.

Electrochemical Microsensor for Microfluidic Glyphosate Monitoring in Water Using MIP-Based Concentrators.

Glyphosate (GLY) is a broad-spectrum herbicide and is the most used pesticide worldwide. This vast usage has raised strong interest in the ecotoxicological impacts and human risks, with contamination of water being a major concern. Decentralized analytical techniques for water monitoring are of high importance. In this work, we present a small, low-cost, and time-effective electrochemical, chip-based microfluidic device for direct electrochemical detection of GLY downstream of a molecularly imprinted polymer (MIP) concentrator. We studied the electrochemical behavior of GLY and its metabolite aminomethylphosphonic acid (AMPA) using cyclic voltammetry with noble metal electrodes in acidic, neutral, and basic media. A chronoamperometric sensor protocol was developed for sensitive and selective GLY measurements on gold electrodes. The optimized protocol was transferred to a chip-based microsensor platform for online and real-time detection of GLY in a microfluidic setup. The results in the range from 0 to 50 µM GLY in 0.5 M H2SO4 show high linearity and a sensitivity of 10.3 ± 0.6 µA mm-2 mM-1 for the chip-based microfluidic platform. Successful recovery of GLY concentrated from untreated tap water and its precise detection from low volumes demonstrates the advantages of our system.

A Real-Time Thermal Sensor System for Quantifying the Inhibitory Effect of Antimicrobial Peptides on Bacterial Adhesion and Biofilm Formation.

The increasing rate of antimicrobial resistance (AMR) in pathogenic bacteria is a global threat to human and veterinary medicine. Beyond antibiotics, antimicrobial peptides (AMPs) might be an alternative to inhibit the growth of bacteria, including AMR pathogens, on different surfaces. Biofilm formation, which starts out as bacterial adhesion, poses additional challenges for antibiotics targeting bacterial cells. The objective of this study was to establish a real-time method for the monitoring of the inhibition of (a) bacterial adhesion to a defined substrate and (b) biofilm formation by AMPs using an innovative thermal sensor. We provide evidence that the thermal sensor enables continuous monitoring of the effect of two potent AMPs, protamine and OH-CATH-30, on surface colonization of bovine mastitis-associated Escherichia (E.) coli and Staphylococcus (S.) aureus. The bacteria were grown under static conditions on the surface of the sensor membrane, on which temperature oscillations generated by a heater structure were detected by an amorphous germanium thermistor. Bacterial adhesion, which was confirmed by white light interferometry, caused a detectable amplitude change and phase shift. To our knowledge, the thermal measurement system has never been used to assess the effect of AMPs on bacterial adhesion in real time before. The system could be used to screen and evaluate bacterial adhesion inhibition of both known and novel AMPs.

CRISPR-powered electrochemical microfluidic multiplexed biosensor for target amplification-free miRNA diagnostics.

Recently the use of microRNAs (miRNAs) as biomarkers for a multitude of diseases has gained substantial significance for clinical as well as point-of-care diagnostics. Amongst other challenges, however, it holds the central requirement that the concentration of a given miRNA must be evaluated within the context of other factors in order to unambiguously diagnose one specific disease. In terms of the development of diagnostic methods and devices, this implies an inevitable demand for multiplexing in order to be able to gauge the abundance of several components of interest in a patient's sample in parallel. In this study, we design and implement different multiplexed versions of our electrochemical microfluidic biosensor by dividing its channel into subsections, creating four novel chip designs for the amplification-free and simultaneous quantification of up to eight miRNAs on the CRISPR-Biosensor X ('X' highlighting the multiplexing aspect of the device). We then use a one-step model assay followed by amperometric readout in combination with a 2-min-stop-flow-protocol to explore the fluidic and mechanical characteristics and limitations of the different versions of the device. The sensor showing the best performance, is subsequently used for the Cas13a-powered proof-of-concept measurement of two miRNAs (miRNA-19b and miRNA-20a) from the miRNA-17-92 cluster, which is dysregulated in the blood of pediatric medulloblastoma patients. Quantification of the latter, alongside simultaneous negative control measurements are accomplished on the same device. We thereby confirm the applicability of our platform to the challenge of amplification-free, parallel detection of multiple nucleic acids.

In Situ Mapping of H2, O2, and H2O2 in Microreactors: A Parallel, Selective Multianalyte Detection Method.

Determining local concentrations of the analytes in state-of-the-art microreactors is essential for the development of optimized and safe processes. However, the selective, parallel monitoring of all relevant reactants and products in a multianalyte environment is challenging. Electrochemical microsensors can provide unique information on the reaction kinetics and overall performance of the hydrogen peroxide synthesis process in microreactors, thanks to their high spatial and temporal resolution and their ability to measure in situ, in contrast to other techniques. We present a chronoamperometric approach which allows the selective detection of the dissolved gases hydrogen and oxygen and their reaction product hydrogen peroxide on the same platinum microelectrode in an aqueous electrolyte. The method enables us to obtain the concentration of each analyte using three specific potentials and to subtract interfering currents from the mixed signal. While hydrogen can be detected independently, no potentials can be found for a direct, selective measurement of oxygen and hydrogen peroxide. Instead, it was found that for combined signals, the individual contribution of all analytes superimposes linearly additive. We showed that the concentrations determined from the subtracted signals correlate very well with results obtained without interfering analytes present. For the first time, this approach allowed the mapping of the distribution of the analytes hydrogen, oxygen, and hydrogen peroxide inside a multiphase membrane microreactor, paving the way for online process control.

On-Site Therapeutic Drug Monitoring.

Recent technological advances have stimulated efforts to bring personalized medicine into practice. Yet, traditional application fields like therapeutic drug monitoring (TDM) have remained rather under-appreciated. Owing to clear dose-response relationships, TDM could improve patient outcomes and reduce healthcare costs. While chromatography-based routine practices are restricted due to high costs and turnaround times, biosensors overcome these limitations by offering on-site analysis. Nevertheless, sensor-based approaches have yet to break through for clinical TDM applications, due to the gap between scientific and clinical communities. We provide a critical overview of current TDM practices, followed by a TDM guideline to establish a common ground across disciplines. Finally, we discuss how the translation of sensor systems for TDM can be facilitated, by highlighting the challenges and opportunities.

Microsensor Electrodes for 3D Inline Process Monitoring in Multiphase Microreactors.

We present an electrochemical microsensor for the monitoring of hydrogen peroxide direct synthesis in a membrane microreactor environment by measuring the hydrogen peroxide and oxygen concentrations. In prior work, for the first time, we performed in situ measurements with electrochemical microsensors in a microreactor setup. However, the sensors used were only able to measure at the bottom of the microchannel. Therefore, only a limited assessment of the gas distribution and concentration change over the reaction channel dimensions was possible because the dissolved gases entered the reactor through a membrane at the top of the channel. In this work, we developed a new fabrication process to allow the sensor wires, with electrodes at the tip, to protrude from the sensor housing into the reactor channel. This enables measurements not only at the channel bottom, but also along the vertical axis within the channel, between the channel wall and membrane. The new sensor design was integrated into a multiphase microreactor and calibrated for oxygen and hydrogen peroxide measurements. The importance of measurements in three dimensions was demonstrated by the detection of strongly increased gas concentrations towards the membrane, in contrast to measurements at the channel bottom. These findings allow a better understanding of the analyte distribution and diffusion processes in the microreactor channel as the basis for process control of the synthesis reaction.

Digital DNA microarray generation on glass substrates.

In this work we show how DNA microarrays can be produced batch wise on standard microscope slides in a fast, easy, reliable and cost-efficient way. Contrary to classical microarray generation, the microarrays are generated via digital solid phase PCR. We have developed a cavity-chip system made of a PDMS/aluminum composite which allows such a solid phase PCR in a scalable and easy to handle manner. For the proof of concept, a DNA pool composed of two different DNA species was used to show that digital PCR is possible in our chips. In addition, we demonstrate that DNA microarray generation can be realized with different laboratory equipment (slide cycler, manually in water baths and with an automated cartridge system). We generated multiple microarrays and analyzed over 13,000 different monoclonal DNA spots to show that there is no significant difference between the used equipment. To show the scalability of our system we also varied the size and number of the cavities located in the array region up to more than 30,000 cavities with a volume of less than 60 pL per cavity. With this method, we present a revolutionary tool for novel DNA microarrays. Together with new established label-free measurement systems, our technology has the potential to give DNA microarray applications a new boost.

A lab-on-a-chip for free-flow electrophoretic preconcentration of viruses and gel electrophoretic DNA extraction.

Nucleic acid amplification techniques such as real-time PCR are essential instruments for the identification and quantification of viruses. They are fast, very sensitive and highly specific, but often require elaborate and labor intensive sample preparation to achieve successful amplification of the target sequence. In this work we demonstrate the complete microfluidic preparation of amplifiable virus DNA from dilute specimens. Our approach combines free-flow electrophoretic preconcentration of viral particles with thermal lysis and gel-electrophoretic nucleic acid extraction on a single device. The on-chip preconcentration achieves a capture efficiency of >99% for dilute suspensions of bacteriophage PhiX174. Following preconcentration, phages are thermally lysed and released DNA is recovered after 40 s of on-chip gel-electrophoresis with a recovery rate of ∼73%. Furthermore we demonstrate a detection limit of ∼1 PFU ml-1 (∼0.02 DNA copies per µl) for the detection of bacteriophage PhiX174 by PCR. To simplify operation of the device, we describe the development of a custom-made chip holder as well as a compact peristaltic pump and power supply, which enable user-friendly operation with low risk of cross-contamination and high potential for automation in the field of point-of-care diagnostics.

Enhanced Protein Immobilization on Polymers-A Plasma Surface Activation Study.

Over the last years, polymers have gained great attention as substrate material, because of the possibility to produce low-cost sensors in a high-throughput manner or for rapid prototyping and the wide variety of polymeric materials available with different features (like transparency, flexibility, stretchability, etc.). For almost all biosensing applications, the interaction between biomolecules (for example, antibodies, proteins or enzymes) and the employed substrate surface is highly important. In order to realize an effective biomolecule immobilization on polymers, different surface activation techniques, including chemical and physical methods, exist. Among them, plasma treatment offers an easy, fast and effective activation of the surfaces by micro/nanotexturing and generating functional groups (including carboxylic acids, amines, esters, aldehydes or hydroxyl groups). Hence, here we present a systematic and comprehensive plasma activation study of various polymeric surfaces by optimizing different parameters, including power, time, substrate temperature and gas composition. Thereby, the highest immobilization efficiency along with a homogenous biomolecule distribution is achieved with a 5-min plasma treatment under a gas composition of 50% oxygen and nitrogen, at a power of 1000 W and a substrate temperature of 80 °C. These results are also confirmed by different surface characterization methods, including SEM, XPS and contact angle measurements.

Impact of assay format on miRNA sensing: Electrochemical microfluidic biosensor for miRNA-197 detection.

MicroRNAs (miRNAs) are important biomarkers for the early detection of various diseases, especially cancer. Therefore, there is a continuing interest in different biosensing strategies that allow for the point-of-care measurement of miRNAs. Almost all miRNA sensors utilize cross-hybridization of the target miRNA with a capture probe for the recognition, which can be designed in either a sandwich or a competitive format. In this work, we present a low-cost microfluidic biosensor platform for the electrochemical measurement of miRNA-197 (a tumor biomarker candidate) in undiluted human serum samples, operating with very low sample volumes (580 nl) and a sample-to-result time of one hour. For this purpose, different on-chip miRNA bioassays based on sandwich and competitive formats are developed and compared in terms of their sensitivity, dynamic range, selectivity, precision, and simplicity. The obtained results show that, despite having a narrower dynamic range when compared to the competitive format, the sandwich assay has superior performance regarding its sensitivity and selectivity. The lowest limit of detection which can be achieved with the sandwich assay is 1.28 nM (0.74 fmole), while 4.05 nM (2.35 fmole) with the competitive format. Moreover, the sandwich assay proves to have a better distinction against single-base mismatch oligonucleotide sequences compared to the competitive one. Due to its versatility and easy handling, overcoming the issue with the sensitivity, the implemented electrochemical microfluidic biosensor could pave the way for rapid and low-cost on-site miRNA diagnostics.