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We adapted an existing, spaceflight-proven, robust "electronic nose" (E-Nose) that uses an array of electrical resistivity-based nanosensors mimicking aspects of mammalian olfaction to conduct on-site, rapid screening for COVID-19 infection by measuring the pattern of sensor responses to volatile organic compounds (VOCs) in exhaled human breath. We built and tested multiple copies of a hand-held prototype E-Nose sensor system, composed of 64 chemically sensitive nanomaterial sensing elements tailored to COVID-19 VOC detection; data acquisition electronics; a smart tablet with software (App) for sensor control, data acquisition and display; and a sampling fixture to capture exhaled breath samples and deliver them to the sensor array inside the E-Nose. The sensing elements detect the combination of VOCs typical in breath at parts-per-billion (ppb) levels, with repeatability of 0.02% and reproducibility of 1.2%; the measurement electronics in the E-Nose provide measurement accuracy and signal-to-noise ratios comparable to benchtop instrumentation. Preliminary clinical testing at Stanford Medicine with 63 participants, their COVID-19-positive or COVID-19-negative status determined by concomitant RT-PCR, discriminated between these two categories of human breath with a 79% correct identification rate using "leave-one-out" training-and-analysis methods. Analyzing the E-Nose response in conjunction with body temperature and other non-invasive symptom screening using advanced machine learning methods, with a much larger database of responses from a wider swath of the population, is expected to provide more accurate on-the-spot answers. Additional clinical testing, design refinement, and a mass manufacturing approach are the main steps toward deploying this technology to rapidly screen for active infection in clinics and hospitals, public and commercial venues, or at home.

Space experiments are a technically challenging but a scientifically important part of astrobiology and astrochemistry research. The International Space Station (ISS) is an excellent example of a highly successful and long-lasting research platform for experiments in space, that has provided a wealth of scientific data over the last two decades. However, future space platforms present new opportunities to conduct experiments with the potential to address key topics in astrobiology and astrochemistry. In this perspective, the European Space Agency (ESA) Topical Team Astrobiology and Astrochemistry (with feedback from the wider scientific community) identifies a number of key topics and summarizes the 2021 "ESA SciSpacE Science Community White Paper" for astrobiology and astrochemistry. We highlight recommendations for the development and implementation of future experiments, discuss types of in situ measurements, experimental parameters, exposure scenarios and orbits, and identify knowledge gaps and how to advance scientific utilization of future space-exposure platforms that are either currently under development or in an advanced planning stage. In addition to the ISS, these platforms include CubeSats and SmallSats, as well as larger platforms such as the Lunar Orbital Gateway. We also provide an outlook for in situ experiments on the Moon and Mars, and welcome new possibilities to support the search for exoplanets and potential biosignatures within and beyond our solar system., (© 2023. The Author(s).)

Small satellite technologies, particularly CubeSats, are enabling breakthrough research in space. Over the past 15 years, NASA Ames Research Center has developed and flown half a dozen biological CubeSats in low Earth orbit (LEO) to conduct space biology and astrobiology research investigating the effects of the space environment on microbiological organisms. These studies of the impacts of radiation and reduced gravity on cellular processes include dose-dependent interactions with antimicrobial drugs, measurements of gene expression and signaling, and assessment of radiation damage. BioSentinel, the newest addition to this series, will be the first deep space biological CubeSat, its heliocentric orbit extending far beyond the radiation-shielded environment of low Earth orbit. BioSentinel's 4U biosensing payload, the first living biology space experiment ever conducted beyond the Earth-Moon system, will use a microbial bioassay to assess repair of radiation-induced DNA damage in eukaryotic cells over a duration of 6-12 months. Part of a special collection of articles focused on BioSentinel and its science mission, this article describes the design, development, and testing of the biosensing payload's microfluidics and optical systems, highlighting improvements relative to previous CubeSat life-support and bioanalytical measurement technologies.

The isomer-specific detection and quantitation of m -, p -, and o -xylene and ethylbenzene, dissolved singly and as mixtures in aqueous solutions at concentrations from 100 to 1200 ppb by volume, is reported for a specifically designed polymer-plasticizer coating on a shear-horizontal surface acoustic wave (SH-SAW) device. The polystyrene-ditridecyl phthalate-blend coating was designed utilizing Hansen solubility parameters and considering the dipole moment and polarizability of the analytical targets and coating components to optimize the affinity of the sensor coating for the four chemical isomers. The two key coating sorption properties, sensitivity and response time constant, are determined by the (slightly different) dipole moments and polarizabilities of the four target analytes: as analyte dipole moment decreases, coating sensitivity increases; as analyte polarizability decreases, coating response time lengthens. Using the measured sensitivities and time constants for the targets, sensor signals were processed with exponentially weighted recursive-least-squares estimation (EW-RLSE) to identify (with near 100% accuracy) and quantify (with ± 5-7% accuracy) the isomers. This impressive performance was achieved by combining the specifically tailored, high-sensitivity coating and an SH-SAW platform (yielding a detection limit of 5 ppb for the analytes) and using the EW-RLS estimator, which estimates unknown parameters accurately even in the presence of measurement noise and for analytes with only minor differences in response. Identification of the xylene isomers is important for applications including environmental monitoring and chemical manufacturing.

A relatively simple design procedure is presented for new, adaptable chemical sensor coatings made from a single polymer - plasticizer pair to detect single or a mixture of chemical compounds (e.g., BTEX, the small aromatic hydrocarbon family). Affinity between coating components and target analytes, expressed through Hansen solubility parameters and relative energy difference values, describes the sensitivity of the resultant coatings to each analyte. While analyte affinity is paramount for plasticizer selection, for the aqueous-phase sensing application described here, it must be traded off with the permanence in the host polymer, i.e., resistance to leaching into the ambient aqueous phase; deleterious effects including coating creep must also be minimized. By varying the polymer:plasticizer mixing ratio, the physical and chemical properties of the resultant coatings can be tuned across a range of sensing properties, in particular the differential response magnitude and rate, for multiple analytes. Together with the measurement of multiple sensor response parameters (relative sensitivity and response time constant) for each coating, this approach allows for identification and quantification of target analytes not previously separable using commercial off-the-shelf (COTS) polymer sensor coatings. Sensing results using a five-sensor array based on five different mixing ratios of a single plasticizer polymer pair (plasticizer: ditridecyl phthalate; polymer: polystyrene) demonstrate unique identification of mixtures of BTEX analytes, including differentiation of the chemical isomers ethylbenzene and total xylene (or "xylenes"), something not previously feasible for separation-free liquid-phase sensing with commercially available polymer coatings. Ultimately, the response of a single optimized sensor coating identified and quantified the components of various mixtures, including identification of likely interferents, using a customized estimation-theory-based multivariate signal-processing technique.

The global COVID-19 pandemic has changed many aspects of daily lives. Wearing personal protective equipment, especially respirators (face masks), has become common for both the public and medical professionals, proving to be effective in preventing spread of the virus. Nevertheless, a detailed understanding of respirator filtration-layer internal structures and their physical configurations is lacking. Here, we report three-dimensional (3D) internal analysis of N95 filtration layers via X-ray tomography. Using deep learning methods, we uncover how the distribution and diameters of fibers within these layers directly affect contaminant particle filtration. The average porosity of the filter layers is found to be 89.1%. Contaminants are more efficiently captured by denser fiber regions, with fibers <1.8 μm in diameter being particularly effective, presumably because of the stronger electric field gradient on smaller diameter fibers. This study provides critical information for further development of N95-type respirators that combine high efficiency with good breathability.

We report the results of the EcAMSat (Escherichia coli Antimicrobial Satellite) autonomous space flight experiment, investigating the role of σ s in the development of antibiotic resistance in uropathogenic E. coli (UPEC) in microgravity (µ-g). The presence of σ s , encoded by the rpoS gene, has been shown to increase antibiotic resistance in Earth gravity, but it was unknown if this effect occurs in µ-g. Two strains, wildtype (WT) UPEC and its isogenic ΔrpoS mutant, were grown to stationary phase aboard EcAMSat, an 11-kg small satellite, and in a parallel ground-based control experiment; cell growth rates for the two strains were found to be unaltered by µ-g. After starvation for over 24 h, stationary-phase cells were incubated with three doses of gentamicin (Gm), a common treatment for urinary tract infections (which have been reported in astronauts). Cellular metabolic activity was measured optically using the redox-based indicator alamarBlue (aB): both strains exhibited slower metabolism in µ-g, consistent with results from previous smallsat missions. The results also showed that µ-g did not enhance UPEC resistance to Gm; in fact, both strains were more susceptible to Gm in µ-g. It was also found, via a second ground-control experiment, that multi-week storage in the payload hardware stressed the cells, potentially obscuring small differential effects of the antibiotic between WT and mutant and/or between µ-g and ground. Overall, results showed that the ∆rpoS mutant was 34-37% less metabolically active than the WT for four different sets of conditions: ground without Gm, ground with Gm; µ-g without Gm, µ-g with Gm. We conclude therefore that the rpoS gene and its downstream products are important therapeutic targets for treating bacterial infections in space, much as they are on the ground., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Published by Elsevier Ltd.)

We report here complete 6-month results from the orbiting Space Environment Survivability of Living Organisms (SESLO) experiment. The world's first and only long-duration live-biology cubesat experiment, SESLO was executed by one of two 10-cm cube-format payloads aboard the 5.5-kg O/OREOS (Organism/Organic Exposure to Orbital Stresses) free-flying nanosatellite, which launched to a 72°-inclination, 650-km Earth orbit in 2010. The SESLO experiment measured the long-term survival, germination, metabolic, and growth responses of Bacillus subtilis spores exposed to microgravity and ionizing radiation including heavy-ion bombardment. A pair of radiation dosimeters (RadFETs, i.e., radiation-sensitive field-effect transistors) within the SESLO payload provided an in-situ dose rate estimate of 6-7.6 mGy/day throughout the mission. Microwells containing samples of dried spores of a wild-type B. subtilis strain and a radiation-sensitive mutant deficient in Non-Homologoous End Joining (NHEJ) were rehydrated after 14, 91, and 181 days in space with nutrient medium containing with the redox dye alamarBlue (aB), which changes color upon reaction with cellular metabolites. Three-color transmitted light intensity measurements of all microwells were telemetered to Earth within days of each 24-hour growth experiment. At 14 and 91 days, spaceflight samples germinated, grew, and metabolized significantly more slowly than matching ground-control samples, as measured both by aB reduction and optical density changes; these rate differences notwithstanding, the final optical density attained was the same in both flight and ground samples. After 181 days in space, spore germination and growth appeared hindered and abnormal. We attribute the differences not to an effect of the space environment per se, as both spaceflight and ground-control samples exhibited the same behavior, but to a pair of ~15-day thermal excursions, after the 91-day measurement and before the 181-day experiment, that peaked above 46 °C in the SESLO payload. Because the payload hardware operated nominally at 181 days, the growth issues point to heat damage, most likely to component(s) of the growth medium (RPMI 1640 containing aB) or to biocompatibility issues caused by heat-accelerated outgassing or leaching of harmful compounds from components of the SESLO hardware and electronics.

Most chemical sensors are only partially selective to any specific target analyte(s), making identification and quantification of analyte mixtures challenging, a problem often addressed using arrays of partially selective sensors. This work presents and experimentally verifies a signal-processing technique based on estimation theory for online identification and quantification of multiple analytes using only the response data collected from a single polymer-coated sensor device. The demonstrated technique, based on multiple stages of exponentially weighted recursive least-squares estimation (EW-RLSE), first determines which of the analytes included in the sensor response model are absent from the mixture being analyzed; these are then eliminated from the model prior to executing the final stage of EW-RLSE, in which the sample's constituent analytes are more accurately quantified. The overall method is based on a sensor response model with specific parameters describing each coating-analyte pair and requires no initial assumptions regarding the concentrations of the analytes in a given sample. The technique was tested using the measured responses of polymer-coated shear-horizontal surface acoustic wave devices to multi-analyte mixtures of benzene, toluene, ethylbenzene, xylenes, and 1,2,4-trimethylbenzene in water. The results demonstrate how this method accurately identifies and quantifies the analytes present in a sample using the measured response of just a single sensor device. This effective, simple, lower-cost alternative to sensor arrays needs no arduous training protocol, just measurement of the response characteristics of each individual target analyte and the likely interferents and/or classes thereof.

Blood type O is associated with a lower risk of myocardial infarction. Platelets play a critical role in myocardial infarction. It is not known whether the expression of blood group antigens on platelet proteins alters platelet function; we hypothesized that platelet function would be different between donors with blood type O and those with non-O. To address this hypothesis, we perfused blood from healthy type O donors (n = 33) or non-O donors (n = 54) over pooled plasma derived von Willebrand factor (VWF) protein and purified blood type-specific VWF at arterial shear and measured platelet translocation dynamics. We demonstrate for the first time that type O platelets travel farther at greater speeds before forming stable bonds with VWF. To further characterize these findings, we used a novel analytical model of platelet interaction. Modeling revealed that the kinetics for GPIb/VWF binding rate are significantly lower for type O compared with non-O platelets. Our results demonstrate that platelets from type O donors interact less with VWF at arterial shear than non-O platelets. Our results suggest a potential mechanism for the reduced risk of myocardial infarction associated with blood type O., (© 2019 by The American Society of Hematology.)

The process of platelet adhesion is initiated by glycoprotein (GP)Ib and GPIIbIIIa receptors on the platelet surface binding with von Willebrand factor on the vascular walls. This initial adhesion and detachment of a single platelet is a complex process that involves multiple bonds forming and breaking and is strongly influenced by the surrounding blood-flow environment. In addition to bond-level kinetics, external factors such as shear rate, hematocrit, and GPIb and GPIIbIIIa receptor densities have also been identified as influencing the platelet-level rate constants in separate studies, but this still leaves a gap in understanding between these two length scales. In this study, we investigate the fundamental relationship of the dynamics of platelet adhesion, including these interrelating factors, using a coherent strategy. We build a, to our knowledge, novel and computationally efficient multiscale model accounting for multibond kinetics and hydrodynamic effects due to the flow of a cellular suspension. The model predictions of platelet-level kinetics are verified by our microfluidic experiments, which systematically investigate the role of each external factor on platelet adhesion in an in vitro setting. We derive quantitative formulas describing how the rates of platelet adhesion, translocation, and detachment are defined by the molecular-level kinetic constants, the local platelet concentration near the reactive surface determined by red-blood-cell migration, the platelet effective reactive area due to its tumbling motion, and the platelet surface receptor density. Furthermore, if any of these aspects involved have abnormalities, e.g., in a disease condition, our findings also have clinical relevance in predicting the resulting change in the adhesion dynamics, which is essential to hemostasis and thrombosis., (Copyright © 2019 Biophysical Society. Published by Elsevier Inc. All rights reserved.)

A new approach is reported to detect and quantify the members of a group of small-aromatic-molecule target analytes: benzene, toluene, ethylbenzene, and xylenes (BTEX), dissolved in water, in the presence of interferents, using only the data collected from a single polymer-coated SH-SAW (shear horizontal surface acoustic wave) device and a two-stage adaptive estimation scheme. This technique is composed of exponentially weighted recursive least-squares estimation (EW-RLSE) and a bank of Kalman filters (BKFs) and does not require any prior knowledge of the initial concentration range of the target analytes. The proposed approach utilizes the transient sensor response to sorption and/or desorption of the analytes as well as the error range associated with the response time constants to provide more information about the analyte-specific interactions with the polymer film. The approach assumes that the sensor response to contaminated groundwater is a linear combination of the responses to the single target analytes, the interferents that interact with the selected polymer sensor coatings, and measurement noise. The proposed technique was tested using actual sensor responses to contaminated groundwater samples containing multiple BTEX compounds with concentrations ranging from 10 to 2000 parts per billion, as well as common interferents including ethanol, 1,2,4-trimethylbenzene, naphthalene, n-heptane, and MTBE (methyl tert-butyl ether). Estimated concentration values, accurate to ±10% for benzene/toluene and ±15% for ethylbenzene/xylenes, are obtained in near-real time. The utilization of sorption and/or desorption data enables detection and quantification of BTEX compounds with improved accuracy, high tolerance to measurement noise, and improved chemical selectivity.

Human immune response is compromised and bacteria can become more antibiotic resistant in space microgravity (MG). We report that under low-shear modeled microgravity (LSMMG), stationary-phase uropathogenic Escherichia coli (UPEC) become more resistant to gentamicin (Gm), and that this increase is dependent on the presence of σ s (a transcription regulator encoded by the rpoS gene). UPEC causes urinary tract infections (UTIs), reported to afflict astronauts; Gm is a standard treatment, so these findings could impact astronaut health. Because LSMMG findings can differ from MG, we report preparations to examine UPEC's Gm sensitivity during spaceflight using the E. coli Anti-Microbial Satellite (EcAMSat) as a free-flying "nanosatellite" in low Earth orbit. Within EcAMSat's payload, a 48-microwell fluidic card contains and supports study of bacterial cultures at constant temperature; optical absorbance changes in cell suspensions are made at three wavelengths for each microwell and a fluid-delivery system provides growth medium and predefined Gm concentrations. Performance characterization is reported here for spaceflight prototypes of this payload system. Using conventional microtiter plates, we show that Alamar Blue (AB) absorbance changes can assess the Gm effect on E. coli viability, permitting telemetric transfer of the spaceflight data to Earth. Laboratory results using payload prototypes are consistent with wellplate and flask findings of differential sensitivity of UPEC and its ∆rpoS strain to Gm. if σ s plays the same role in space MG as in LSMMG and Earth gravity, countermeasures discovered in recent Earth studies (aimed at weakening the UPEC antioxidant defense) to control UPEC infections would prove useful also in space flights. Further, EcAMSat results should clarify inconsistencies from previous space experiments on bacterial antibiotic sensitivity and other issues., (Copyright © 2017. Published by Elsevier Ltd.)

Platelet function in pregnancy is poorly understood. Previous studies of platelet function in pregnancy have used non-physiological assays of platelet function with conflicting results. This study using a physiological assay of platelet function investigated platelet interactions with von Willebrand Factor (VWF) in blood from healthy pregnant women and healthy non-pregnant controls. Blood samples (200 µl) from third-trimester pregnancies (n = 21) and non-pregnant controls (n = 21) were perfused through custom-made parallel-plate flow chambers coated with VWF under arterial shear (1,500 s -1 ). Multi-parameter measurements of platelet interactions with the immobilized VWF surface were recorded by digital-image microscopy and analysed using custom-designed platelet-tracking software. Platelet interactions with VWF decreased in healthy third-trimester pregnant participants relative to controls. This effect is most likely due to haemodilution which occurs physiologically during pregnancy. Interestingly, platelets in blood from pregnant participants translocated more slowly on VWF under arterial-shear conditions. These decreases in platelet translocation speed were independent of haemodilution, suggesting intrinsic changes in platelet function with pregnancy.

We report the first-ever direct detection of benzene in water at concentrations below 100 ppb (parts per billion) using acoustic wave (specifically, shear-horizontal surface acoustic wave, SH-SAW) sensors with plasticized polymer coatings. Two polymers and two plasticizers were studied as materials for sensor coatings. For each polymer-plasticizer combination, the influence of the mixing ratio of the blend on the sensitivity to benzene was measured and compared to commercially available polymers that were used for BTEX (benzene, toluene, ethylbenzene, and xylene) detection in previous work. After optimizing the coating parameters, the highest sensitivity and lowest detection limit for benzene were found for a 1.25 μm thick sensor coating of 17.5%-by-weight diisooctyl azelate-polystyrene on the tested acoustic wave device. The calculated detection limit was 45 ppb, with actual sensor responses to concentrations down to 65 ppb measured directly. Among the sensor coatings that showed good sensitivity to benzene, the best long-term stability was found for a 1.0 μm thick coating of 23% diisononyl cyclohexane-1,2-dicarboxylate-polystyrene, which was studied here because it is known to show no detectable leaching in water. The present work demonstrates that, by varying type of plasticizer, mixing ratio, and coating thickness, the mechanical and chemical properties of the coatings can be conveniently tailored to maximize analyte sorption and partial chemical selectivity for a given class of analytes as well as to minimize acoustic-wave attenuation in contact with an aqueous phase at the operating frequency of the sensor device.

The imaging of shear-mediated dynamic platelet behavior interacting with surface-immobilized von Willebrand factor (vWF) has tremendous potential in characterizing changes in platelet function for clinical diagnostics purposes. However, the imaging output, a series of images representing platelets adhering and rolling on the surface, poses unique, non-trivial challenges for software algorithms that reconstruct the positional trajectories of platelets. We report on an algorithm that tracks platelets using the output of such flow run experiments, taking into account common artifacts encountered by previously-published methods, and we derive seven key metrics of platelet dynamics that can be used to characterize platelet function. Extensive testing of our method using simulated platelet flow run data was carried out to validate our tracking method and derived metrics in capturing key platelet-vWF interaction-dynamics properties. Our results show that while the number of platelets present on the imaged area is the leading cause of errors, flow run data from two experiments using whole blood samples showed that our method and metrics can detect platelet property changes/differences that are concordant with the expected biological outcome, such as inhibiting key platelet receptors such as P2Y 1 , glycoprotein (GP)Ib and GPIIb/IIIa. These findings support the use of our methodologies to characterize platelet function among a wide range of healthy and disease cohorts.

While critically important, the platelet function at the high shear rates typical of the microcirculation is relatively poorly understood. Using a large scale Stokes flow simulation, Zhao et al. recently showed that RBC-induced velocity fluctuations cause platelets to marginate into the RBC free near-wall region [Zhao et al., Physics of Fluids, 2012, 24, 011902]. We extend their work by investigating the dynamics of platelets in shear after margination. An overall platelet adhesion model is proposed in terms of a continuous time Markov process and the transition rates are established with numerical simulations involving platelet-wall adhesion. Hydrodynamic drag and Brownian forces are calculated with the boundary element method, while the RBC collisions are incorporated through an autoregressive process. Hookean springs with first order bond kinetics are used to model receptor-ligand bonds formed between the platelet and the wall. The simulations are compared with in vitro microfluidic experiments involving platelet adhesion to Von Willebrand Factor (VWF) coated surfaces.

In this paper we describe a method for three-dimensional wax patterning of microfluidic paper-based analytical devices (μPADs). The method is rooted in the fundamental details of wax transport in paper and provides a simple way to fabricate complex channel architectures such as hemichannels and fully enclosed channels. We show that three-dimensional μPADs can be fabricated with half as much paper by using hemichannels rather than ordinary open channels. We also provide evidence that fully enclosed channels are efficiently isolated from the exterior environment, decreasing contamination risks, simplifying the handling of the device, and slowing evaporation of solvents.

We report the development and demonstration of an assay that distinguishes the pharmacological effects of two widely used antiplatelet therapies, aspirin (COX-1 inhibitor) and clopidogrel (P2Y12 inhibitor). Whole blood is perfused through a low-volume microfluidic device in contact with a well-characterized (ellipsometry, atomic force microscopy) acid-soluble type I collagen surface. Whole human blood treated in vitro with a P2Y12 inhibitor 2-methylthioadenosine 5'-monophosphate triethylammonium salt (2-MeSAMP) extended the time to the start of platelet recruitment, i.e., platelet binding to the collagen surface. Treatment with 2-MeSAMP also slowed the rate of aggregate buildup, with an overall reduced average platelet aggregate area after 8 min of constant blood flow. A far smaller effect was observed for in vitro treatment with aspirin, for which the rate of change of surface coverage is indistinguishable from controls. In whole blood obtained from patients under treatment with dual-antiplatelet therapy (aspirin and clopidogrel), a significant extension of time to platelet recruitment was observed along with a slowed rate of aggregate buildup and an average aggregate size approximately half that of control measurements. Differentiation of the pharmacological effects of these two well-targeted antiplatelet pathways suggests a role for this assay in determining the antiplatelet effects of these and related new therapeutics in clinical settings.

Lab-on-a-chip (LOC) applications in environmental, biomedical, agricultural, biological, and spaceflight research require an ion-selective electrode (ISE) that can withstand prolonged storage in complex biological media (1-4). An all-solid-state ion-selective-electrode (ASSISE) is especially attractive for the aforementioned applications. The electrode should have the following favorable characteristics: easy construction, low maintenance, and (potential for) miniaturization, allowing for batch processing. A microfabricated ASSISE intended for quantifying H(+), Ca(2+), and CO3(2-) ions was constructed. It consists of a noble-metal electrode layer (i.e. Pt), a transduction layer, and an ion-selective membrane (ISM) layer. The transduction layer functions to transduce the concentration-dependent chemical potential of the ion-selective membrane into a measurable electrical signal. The lifetime of an ASSISE is found to depend on maintaining the potential at the conductive layer/membrane interface (5-7). To extend the ASSISE working lifetime and thereby maintain stable potentials at the interfacial layers, we utilized the conductive polymer (CP) poly(3,4-ethylenedioxythiophene) (PEDOT) (7-9) in place of silver/silver chloride (Ag/AgCl) as the transducer layer. We constructed the ASSISE in a lab-on-a-chip format, which we called the multi-analyte biochip (MAB) (Figure 1). Calibrations in test solutions demonstrated that the MAB can monitor pH (operational range pH 4-9), CO3(2-) (measured range 0.01 mM - 1 mM), and Ca(2+) (log-linear range 0.01 mM to 1 mM). The MAB for pH provides a near-Nernstian slope response after almost one month storage in algal medium. The carbonate biochips show a potentiometric profile similar to that of a conventional ion-selective electrode. Physiological measurements were employed to monitor biological activity of the model system, the microalga Chlorella vulgaris. The MAB conveys an advantage in size, versatility, and multiplexed analyte sensing capability, making it applicable to many confined monitoring situations, on Earth or in space. Biochip Design and Experimental Methods The biochip is 10 x 11 mm in dimension and has 9 ASSISEs designated as working electrodes (WEs) and 5 Ag/AgCl reference electrodes (REs). Each working electrode (WE) is 240 μm in diameter and is equally spaced at 1.4 mm from the REs, which are 480 μm in diameter. These electrodes are connected to electrical contact pads with a dimension of 0.5 mm x 0.5 mm. The schematic is shown in Figure 2. Cyclic voltammetry (CV) and galvanostatic deposition methods are used to electropolymerize the PEDOT films using a Bioanalytical Systems Inc. (BASI) C3 cell stand (Figure 3). The counter-ion for the PEDOT film is tailored to suit the analyte ion of interest. A PEDOT with poly(styrenesulfonate) counter ion (PEDOT/PSS) is utilized for H(+) and CO3(2-), while one with sulphate (added to the solution as CaSO4) is utilized for Ca(2+). The electrochemical properties of the PEDOT-coated WE is analyzed using CVs in redox-active solution (i.e. 2 mM potassium ferricyanide (K3Fe(CN)6)). Based on the CV profile, Randles-Sevcik analysis was used to determine the effective surface area (10). Spin-coating at 1,500 rpm is used to cast ~2 μm thick ion-selective membranes (ISMs) on the MAB working electrodes (WEs). The MAB is contained in a microfluidic flow-cell chamber filled with a 150 μl volume of algal medium; the contact pads are electrically connected to the BASI system (Figure 4). The photosynthetic activity of Chlorella vulgaris is monitored in ambient light and dark conditions.

We present the design and performance characteristics of a platelet analysis platform based on a microfluidic impedance cytometer. Dielectrophoretic focusing is used to centre cells in a fluid stream, which then forms the core of a two-phase flow (dielectric focusing). This flow then passes between electrodes for analysis by differential impedance spectroscopy at multiple frequencies from 280 kHz to 4 MHz. This approach increases the signal-to-noise ratio relative to a single-phase, unfocused stream, while minimising the shear forces to which the cells are subjected. The percentage of activated platelets before and after passage through the chip was measured using flow cytometry, and no significant change was measured. Measuring the in-phase amplitude at a single frequency is sufficient to distinguish platelets from erythrocytes. Using multi-frequency impedance measurements and discriminant analysis, resting platelets can be discriminated from activated platelets. This multifrequency impedance cytometer therefore allows ready determination of the degree of platelet activation in blood samples.

We report a novel device to analyze cell-surface interactions under controlled fluid-shear conditions on well-characterised protein surfaces. Its performance is demonstrated by studying platelets interacting with immobilised von Willebrand Factor at arterial vascular shear rates using just 200 μL of whole human blood per assay. The device's parallel-plate flow chamber, with 0.1 mm² cross sectional area and height-to-width ratio of 1:40, provides uniform, well-defined shear rates along the chip surface with negligible vertical wall effects on the fluid flow profile while minimizing sample volumetric flow. A coating process was demonstrated by ellipsometry, atomic force microscopy, and fluorescent immunostaining to provide reproducible, homogeneous, uniform protein layers over the 0.7 cm² cell-surface interaction area. Customized image processing quantifies dynamic cellular surface coverage vs. time throughout the whole-blood-flow assay for a given drug treatment or disease state. This device can track the dose response of anti-platelet drugs, is suitable for point-of-care diagnostics, and is designed for adaptation to mass manufacture.

We report a rapid, low-cost, portable microfluidic sedimentation cytometer (SeCy) for assessing the somatic cell count and fat content of milk in 15 min using a "sample-in, answer-out" approach. The system consists of 12 independent microfluidic devices, essentially flattened funnel structures, fabricated on the footprint of a single plastic compact disc (CD). Each funnel structure holds 150 μL of milk, has an inlet for milk filling and an outlet for air to escape, and ends in a narrow, closed-end microfluidic channel that facilitates packing of the cells into a column whose length is proportional to cell count. The closed-end channel provides accurate cell counts over the range 50,000->3,000,000 cells per mL. The assay separates cells and fat globules based on their densities (by differential sedimentation), concentrating white cells in the closed-end channel near the outer rim of the CD for estimation of total "cell pellet" volume, while fat globules move toward the center of disc rotation, forming a fat "band" in the funnel. After adding milk to two or more microfluidic devices, the CD is loaded onto a custom-built reader unit that spins the disc for 15 min. Two low-cost microscopes in the reader image the centrifuged cell pellet and the fat band, providing a sufficiently accurate cell count to diagnose mastitis and measuring fat content as an indication of health and nutritional status.

We report an integrated platelet translocation analysis system that measures complex dynamic platelet-protein surface interactions in microliter volumes of unmodified anticoagulated whole blood under controlled fluid shear conditions. The integrated system combines customized platelet-tracking image analysis with a custom-designed microfluidic parallel plate flow chamber and defined von Willebrand factor surfaces to assess platelet trajectories. Using a position-based probability function that accounts for image noise and preference for downstream movement, outputs include instantaneous and mean platelet velocities, periods of motion and stasis, and bond dissociation kinetics. Whole blood flow data from healthy donors at an arterial shear rate (1500 s(-1)) show mean platelet velocities from 8.9+/-1.0 to 12+/-4 microm s(-1). Platelets in blood treated with the antiplatelet agent c7E-Fab fragment spend more than twice as much time in motion as platelets from untreated control blood; the bond dissociation rate constant (k(off)) increases 1.3-fold, whereas mean translocation velocities do not differ. Blood from healthy unmedicated donors was used to assess flow assay reproducibility, donor variability, and the effects of antiplatelet treatment. This integrated system enables reliable, rapid populational quantification of platelet translocation under pathophysiological vascular fluid shear using as little as 150 microl of blood., (Copyright 2010 Elsevier Inc. All rights reserved.)