New highly sensitive and selective catalytic DNA biosensors for metal ions

While remarkable progress has been made in developing sensors for metal ions such as Ca(II) and Zn(II), designing and synthesizing sensitive and selective metal ion sensors remains a significant challenge. Perhaps the biggest challenge is the design and synthesis of a sensor capable of specific and strong metal binding. Since our knowledge about the construction of metal-binding sites in general is limited, searching for sensors in a combinatorial way is of significant value. Therefore, we have been able to use a combinatorial method called in vitro selection to obtain catalytic DNA that can bind a metal ion of choice strongly and specifically. The metal ion selectivity of the catalytic DNA was further improved using a 'negative selection' strategy where catalytic DNA that are selective for competing metal ions are discarded in the in vitro selection processes. By labeling the resulting catalytic DNA with a fluorophore/quencher pair, we have made a new class of metal ion fluorescent sensors that are the first examples of catalytic DNA biosensors for metal ions. The sensors combine the high selectivity of catalytic DNA with the high sensitivity of fluorescent detection, and can be applied to the quantitative detection of metal ions over a wide concentration range and with high selectivity. The use of DNA sensors in detection and quantification of lead ions in environmental samples such as water from Lake Michigan has been demonstrated. DNA is stable, cost-effective, environmentally benign, and easily adaptable to optical fiber and microarray technology for device manufacture. Thus, the DNA sensors explained here hold great promise for on-site and real-time monitoring of metal ions in the fields of environmental monitoring, developmental biology, clinical toxicology, wastewater treatment, and industrial process monitoring.     

Real-time monitoring of the strand displacement amplification (SDA) of human cytomegalovirus by a new SDA-piezoelectric DNA sensor system

Nucleic acid amplification has long been used in biosensor technologies, such as DNA sensors, DNA chips and microarrays, due to its advantage of high sensitivity in detecting target DNA. However, dynamic monitoring of nucleic acid amplifications with traditional DNA sensors in real-time is difficult since a constant temperature must be maintained during detection. Thus, the piezoelectric sensor, one type of traditional DNA sensor, is not applicable in real-time monitoring PCR due to the dramatic change in temperature that occurs during reaction. In this study, we introduced strand displacement amplification (SDA), an well-developed nucleic acid amplification technique that can work under conditions of constant temperature, into the development of a novel piezoelectric sensor. Using the new SDA-piezoelectric DNA sensor, we designed a stable system for liquid-phase detection, in which the crystal oscillator plate was fixed by an easily adjustable screw-threaded clamping mechanism and successfully applied the new sensor system to real-time SDA monitoring of human cytomegalovirus (HCMV). This new technique overcomes the shortcomings of traditional DNA sensors in real-time monitoring of nucleic acid amplification. The technique has proved to be a markedly simplified procedure with a number of advantages, such as higher sensitivity, better time efficiency, and the ability of dynamic real-time detection.     

Developing sensors for real-time measurement of high Ca2+ concentrations

Ca2+ regulates numerous biological processes through spatiotemporal changes in the cytosolic Ca2+ concentration and subsequent interactions with Ca2+ binding proteins. The endoplasmic reticulum (ER) serves as an intracellular Ca2+ store and plays an essential role in cytosolic Ca2+ homeostasis. There is a strong need to develop Ca2+ sensors capable of real-time quantitative Ca2+ concentration measurements in specific subcellular environments without using natural Ca2+ binding proteins such as calmodulin, which themselves participate as signaling molecules in cells. In this report, a strategy for creating such sensors by grafting a Ca2+-binding motif into chromophore sensitive locations in green fluorescence protein is described. The engineered Ca2+ sensors exhibit large ratiometric fluorescence and absorbance changes upon Ca2+ binding with affinities corresponding to the Ca2+ concentrations found in the ER (Kd values range from 0.4 to 2 mM). In addition to characterizing the optical and metal binding properties of the newly developed Ca2+ sensors with various spectroscopic methods, we also examined the kinetic properties using stopped-flow spectrofluorimetry to ensure accurate monitoring of dynamic Ca2+ changes. The developed Ca2+ sensor was successfully targeted to the ER of mammalian cell lines to monitor Ca2+ changes occurring in this compartment in response to stimulation with agonists. We envision that this class of Ca2+ sensors can be modified further to measure the Ca2+ concentration in other cellular compartments, providing tools for studying the contribution of these compartments to cellular Ca2+ signaling.     

Centrale de mesure ambulatoire biomédicale sur PSoC

Improvement in quality and efficiency of health and medicine, at home and in hospital, has become of paramount importance. The solution to this problem would require the continuous monitoring of several key patient parameters, including the assessment of autonomic nervous system (ANS) activity using non-invasive sensors, providing information for emotional, sensorial, cognitive and physiological analysis of the patient. Recent advances in embedded systems, microelectronics, sensors and wireless networking enable the design of wearable systems capable of such advanced health monitoring. The subject of this article is an ambulatory system comprising of a small wrist device connected to several sensors for the detection of the autonomic nervous system activity. It affords monitoring of skin resistance, skin temperature and heart activity. It is also capable of recording the data on a removable media or sending it to computer via a wireless communication. The wrist device is based on a programmable system-on-chip (PSoC) from Cypress.     

Biosensors for real-time in vivo measurements

The current status of sensors capable of continuous measurement of analytes in biological media is reviewed. This review containing 173 references deals with devices whose use in single cells, tissue slices, animal models and humans has been demonstrated. In addition to sensors specific for glucose, lactate, glutamate, pyruvate, choline and acetylcholine, insights obtained from monitoring nitric oxide, Na(+), K(+), Ca(2+), and dopamine are presented. Performance criteria for sensor performance are described as is the subject of biosensor calibration. Biocompatibility issues are dealt with in some detail as is the status of continuous blood glucose monitoring in humans.     

A novel analytical method for in vivo phosphate tracking

Genetically-encoded fluorescence resonance energy transfer (FRET) sensors for phosphate (P(i)) (FLIPPi) were engineered by fusing a predicted Synechococcus phosphate-binding protein (PiBP) to eCFP and Venus. Purified fluorescent indicator protein for inorganic phosphate (FLIPPi), in which the fluorophores are attached to the same PiBP lobe, shows P(i)-dependent increases in FRET efficiency. FLIPPi affinity mutants cover P(i) changes over eight orders of magnitude. COS-7 cells co-expressing a low-affinity FLIPPi and a Na(+)/P(i) co-transporter exhibited FRET changes when perfused with 100 microM P(i), demonstrating concentrative P(i) uptake by PiT2. FLIPPi sensors are suitable for real-time monitoring of P(i) metabolism in living cells, providing a new tool for fluxomics, analysis of pathophysiology or changes of P(i) during cell migration.     

Long-term <Emphasis Type="Italic">ex vivo</Emphasis> monitoring of <Emphasis Type="Italic">in vivo</Emphasis> microRNA activity in liver using a secreted luciferase sensor

Technology for monitoring in vivo microRNA (miRNA) activity is extremely important for elucidating miRNA biology. However, in vivo studies of miRNA have been hampered by the lack of a convenient approach to reliably reflect real-time functional changes in miRNAs. Sensors for miRNA were developed by adding miRNA target sequences to the 3′-untranslated region of Gaussia princeps luciferase (Gluc) mRNA. These sensors were then evaluated in vitro and in vivo by measuring Gluc activity in cell supernatants and in peripheral blood. Sensors driven by the CMV promoter were effective for monitoring miR-122 in living cells, but not for the long-term monitoring of miR-122 or miR-142 in mouse liver because of CMV-promoter silencing. Replacing the CMV promoter with a CAG promoter rendered these sensors effective for the long-term monitoring of relevant liver miRNA activities. We subsequently used the CAG-promoter-based sensor for the long-term monitoring of endogenous liver miR-122, miR142 and miR-34a activities, as well as for exogenous miR-34a activity. Our study demonstrates that real-time in vivo activities of miRNAs can be continuously and conveniently detected in mouse liver using the sensors that we have developed.     

Technology outlook for real-time quality attribute and process parameter monitoring in biopharmaceutical development—A review

Real-time monitoring of bioprocesses by the integration of analytics at critical unit operations is one of the paramount necessities for quality by design manufacturing and real-time release (RTR) of biopharmaceuticals. A well-defined process analytical technology (PAT) roadmap enables the monitoring of critical process parameters and quality attributes at appropriate unit operations to develop an analytical paradigm that is capable of providing real-time data. We believe a comprehensive PAT roadmap should entail not only integration of analytical tools into the bioprocess but also should address automated-data piping, analysis, aggregation, visualization, and smart utility of data for advanced-data analytics such as machine and deep learning for holistic process understanding. In this review, we discuss a broad spectrum of PAT technologies spanning from vibrational spectroscopy, multivariate data analysis, multiattribute chromatography, mass spectrometry, sensors, and automated-sampling technologies. We also provide insights, based on our experience in clinical and commercial manufacturing, into data automation, data visualization, and smart utility of data for advanced-analytics in PAT. This review is catered for a broad audience, including those new to the field to those well versed in applying these technologies. The article is also intended to give some insight into the strategies we have undertaken to implement PAT tools in biologics process development with the vision of realizing RTR testing in biomanufacturing and to meet regulatory expectations.     

Tracking cancer cell proliferation on a CMOS capacitance sensor chip

We report a novel technique for assessing cell proliferation that employs integrated capacitance sensors for monitoring the growth of anchorage-dependent living cells. The sensors measure substrate coupling capacitances of cells cultured on-chip in a standard in vitro environment. The biophysical phenomenon underlying the capacitive behavior of cells is the counterionic polarization around the insulating cell bodies when exposed to weak, low frequency electric fields. The sensors employ charge sharing for mapping sensed capacitance values in the fF range to output voltage signals. The sensor chip has been fabricated in a commercially available 0.5microm, 2-poly 3-metal CMOS technology. We report experimental results demonstrating sensor response to the adhesion of MDA-MB-231 breast cancer cells followed by their proliferation on the chip surface. On-chip capacitance sensing offers a non-invasive, label-free, easy-to-use, miniaturized technique with real-time monitoring capability for tracking cell proliferation in vitro.     

An automated system for biocide testing on biofilms*

The paper presents and discusses a novel on-line real-time non-destructive continuous-flow system for biocide testing on industrial biofilms. This laboratory system is capable of monitoring changes in growth, accumulation and respiratory activity of biofilms in response to biocidal treatment. The system incorporates a fouling monitor for continuous measuring of the rate of biofilm accumulation (heat transfer resistance), a sensor for monitoring of microbial activity (oxygen meter for monitoring the rate of biofilm respiratory activity), and subsystems necessary for microbial life support and control of operation parameters. Examples of system operation and testing of oxidizing and non-oxidizing biocides are presented. Received 25 May 1997/ Accepted in revised form 25 November 1997     

Surface Plasmon Resonance Imaging Sensors: A Review

Surface plasmon resonance (SPR) imaging sensors realize label-free, real-time, highly sensitive, quantitative, high-throughput biological interaction monitoring and the binding profiles from multi-analytes further provide the binding kinetic parameters between different biomolecules. In the past two decades, SPR imaging sensors found rapid increasing applications in fundamental biological studies, medical diagnostics, drug discovery, food safety, precision measurement, and environmental monitoring. In this paper, we review the recent advances of SPR imaging sensor technology towards high-throughput multi-analyte screening. Finally, we describe our multiplex spectral-phase SPR imaging biosensor for high-throughput biosensing applications.     

Designing a Non-invasive Surface Acoustic Resonator for Ultra-high Sensitive Ethanol Detection for an On-the-spot Health Monitoring System

Surface acoustic wave (SAW) sensors–based on piezoelectric crystal resonators–are extremely sensitive to even very small perturbations in the external atmosphere, because the energy associated with the acoustic waves is confined to the crystal surface. In this study, we present a critical review of the recent researches and developments predominantly used for SAW-based organic vapor sensors, especially ethanol. Besides highlighting their potential to realize real-time ethanol sensing, their drawbacks such as indirect sensing, invasive, time initializing, and low reliability, are properly discussed. The study investigates a proposed YZ-lithium niobate piezoelectric substrate with interdigital transducers patterned on the surface. Design of the resonator plays an important role in improving mass sensitivity, particularly the sensing area. Accordingly, a tin dioxide (SnO₂) layer with a specific thickness is generated on the surface of the sensor because of its high affinity to ethanol molecules. To determine the values of sensor configuration without facing the practical problems and the long theoretical calculation time, it is shown that the mass sensitivity of SAW sensors can be calculated by a simple three-dimensional (3-D) finite element analysis (FEA) using a commercial finite-element platform. In design validation step, different concentrations of ethanol are applied to investigate the acoustic wave properties of the sensor. The FEA data are used to obtain the surface and bulk total displacements of the sensor and fast Fourier transform (FFT) on output spectrum. The sensor could develop into highly sensitive and fast responsive structure so that a positive intensity shift of 0.18e-2 RIU is observed when the sensor is exposed to 15 ppm ethanol. It is capable of continuously monitoring the ethanol gas whether as an ultra-high sensitive sensor or switching applications for medical and industrial purposes.     

On-line soft sensing in upstream bioprocessing

This review provides an overview and a critical discussion of novel possibilities of applying soft sensors for on-line monitoring and control of industrial bioprocesses. Focus is on bio-product formation in the upstream process but also the integration with other parts of the process is addressed. The term soft sensor is used for the combination of analytical hardware data (from sensors, analytical devices, instruments and actuators) with mathematical models that create new real-time information about the process. In particular, the review assesses these possibilities from an industrial perspective, including sensor performance, information value and production economy. The capabilities of existing analytical on-line techniques are scrutinized in view of their usefulness in soft sensor setups and in relation to typical needs in bioprocessing in general. The review concludes with specific recommendations for further development of soft sensors for the monitoring and control of upstream bioprocessing.     

Childhood craniopharyngioma. Recent advances in diagnosis, treatment and follow-up

The increased availability of continuous glucose sensors is likely to have a significant impact on pediatric diabetes therapy and education in the future. While a recent meta-analysis of retrospective first-generation Holter-type sensors was not able to show advantages compared to self-monitoring of blood glucose levels, this review focuses on current real-time sensors. Our own experience with 23 children aged 10 (3-15) years with sensor-augmented pump therapy showed high ratings for overall satisfaction and ease of use of the system (5.8 on a 7 point Likert scale). The results of our ongoing international pediatric ONSET trial will show the impact of this technology on learning to live with diabetes by having the biofeedback of glucose excursions from the onset of diabetes. Families previously relying on self-monitoring of blood glucose need to understand the difference between estimating the absolute blood glucose value (point accuracy) and the change in blood glucose (rate accuracy), and how to take into consideration the inherent interstitial time lag. Selection of patients capable and motivated to use continuous sensors accompanied with proper age-appropriate education remain key factors for the long-term success of these new technological advances in diabetes therapy as long as closed loop systems are not available.     

Monitoring blood coagulation with magnetoelastic sensors

The determination of blood coagulation time is an essential part of monitoring therapeutic anticoagulants. Standard methodologies for the measurement of blood clotting time require dedicated personnel and involve blood sampling procedures. A new method based on magnetoelastic sensors has been employed for the monitoring of blood coagulation. The ribbon-like magnetoelastic sensor oscillates at a fundamental frequency, which shifts linearly in response to applied mass loads or a fixed mass load of changing elasticity. The magnetoelastic sensors emit magnetic flux, which can be detected by a remotely located pick-up coil, so that no direct physical connections are required. During blood coagulation, the viscosity of blood changes due to the formation of a soft fibrin clot. In turn, this change in viscosity shifts the characteristic resonance frequency of the magnetoelastic sensor enabling real-time continuous monitoring of this biological event. By monitoring the signal output as a function of time, a distinct blood clotting profile can be seen. The relatively low cost of the magnetoelastic ribbons enables their use as disposable sensors. This, along with the reduced volume of blood required, make the magnetoelastic sensors well suited for at-home and point-of-care testing devices.     

Sensor systems,electronic tongues and electronic noses,for the monitoring of biotechnological processes

Production of biofuel is based on the conversion by microorganisms of complex organic substrates into the methane or ethanol, which are consequently used as energy sources. Real time monitoring of the fermented media composition is of paramount for the effectiveness of the whole process. However, despite the fact that products worth billions of dollars are produced through fermentation processes annually, analytical instruments used for these processes’ monitoring remain relatively primitive. Established laboratory techniques produce exhaustive information about media composition but analysis is often quite time-consuming, expensive, requires skilled personnel and hardly can be automated. Lack of on-line sensors for the fermentation monitoring is commonly stressed in the literature. One of the techniques particularly suitable for this purpose is chemical sensors. Such features as low prices, relatively simple instrumentation, minimal sample preparation and easy automation of measurements make chemical sensors an attractive tool for industrial process control. However, practical use of chemical sensors in complex media is often hindered by their insufficient selectivity. For example, only pH and oxygen probes are routinely used in bio-reactors. One of the emerging approaches permitting to overcome the selectivity problems is the use of systems instead of discrete sensors. Such systems for liquid and gas analysis were named electronic tongues and electronic noses correspondingly. They are capable to perform both quantitative analysis (components’ concentrations) and classification or recognition of multicomponent media. This review presents recent achievements in the R&D and applications of electronic tongues and noses to the monitoring of biotechnological processes. JIMB-2008: BioEnergy—Special issue.     

On-line infrared spectroscopy for bioprocess monitoring

One of the major aims of bioprocess engineering is the real-time monitoring of important process variables. This is the basis of precise process control and is essential for high productivity as well as the exact documentation of the overall production process. Infrared spectroscopy is a powerful analytical technique to analyze a wide variety of organic compounds. Thus, infrared sensors are ideal instruments for bioprocess monitoring. The sensors are non-invasive, have no time delay due to sensor response times, and have no influence on the bioprocess itself. No sampling is necessary, and several components can be analyzed simultaneously. In general, the direct monitoring of substrates, products, metabolites, as well as the biomass itself is possible. In this review article, insights are provided into the different applications of infrared spectroscopy for bioprocess monitoring and the complex data interpretation. Different analytical techniques are presented as well as example applications in different areas.     

Monitoring batch fermentations with an electronic tongue

An electronic tongue comprising 21 potentiometric chemical sensors with pattern recognition tools was used for the rapid off-line monitoring of batch Escherichia coli fermentations. The electronic tongue was capable of monitoring the changes in the media composition as the fermentation progressed, and could correlate this with an increase in biomass. The electronic tongue was also able to monitor the increase in organic acids, especially acetic acid, throughout the fermentation. This technique clearly shows promise as a rapid tool for fermentation monitoring.     

Feasibility of active handheld NDVI sensors for monitoring lichen ground cover

Vegetation indices are corner stones in vegetation monitoring. However, previous field studies on lichens and NDVI have been based on passive sensors. Active handheld sensors, with their own light sources, enables high-precision monitoring under variable ambient conditions. We investigated the use of handheld sensor NDVI for monitoring pale lichen cover across three study sites from boreal heathlands to High Arctic tundra (62–79 °N), and compared it with Sentinel-2 satellite NDVI. NDVI decreased with increasing cover of pale lichens but the correlation between active and satellite NDVI varied between areas. NDVI values declined with lichen cover and ranged from 0.4–0.18 when lichen cover was above 40%. Active ground measurements of NDVI explained 81% of the variation in the satellite NDVI values in Svalbard (High Arctic), while the relationships were lower (∼30% explained variation) in boreal regions (Troms-Finnmark and Røros). We show that active sensors are feasible for extracting information from lichen-dominated vegetation.     

Toward continuous glucose monitoring with planar modified biosensors and microdialysis. Study of temperature, oxygen dependence and in vivo experiment

Glucose biosensors based on the use of planar screen-printed electrodes modified with an electrochemical mediator and with glucose oxidase have been optimised for their application in the continuous glucose monitoring in diabetic patients. A full study of their operative stability and temperature dependence has been accomplished, thus giving useful information for in vivo applications. The effect of dissolved oxygen concentration in the working solution was also studied in order to evaluate its effect on the linearity of the sensors. Glucose monitoring performed with serum samples was performed to evaluate the effect of matrix components on operative stability and demonstrated an efficient behaviour for 72 h of continuous monitoring. Finally, these studies led to a sensor capable of detecting glucose at concentrations as low as 0.04 mM and with a good linearity up to 2.0 mM (at 37 degrees C) with an operative stability of ca. 72 h, thus demonstrating the possible application of these sensors for continuous glucose monitoring in conjunction with a microdialysis probe. Moreover, preliminary in vivo experiments for ca. 20 h have demonstrated the feasibility of this system.