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The design of electrode interfaces to achieve efficient electron transfer to biomolecules is important in many bioelectrochemical processes. Within the field of biohybrid solar energy conversion, constructing multilayered Photosystem I (PSI) protein films that maintain good electronic connection to an underlying electrode has been a major challenge. Previous shortcomings include low loadings, long deposition times, and poor connection between PSI and conducting polymers within composite films. Here, we show that PSI protein complexes can be deposited into monolayers within a 30 min timespan by leveraging the electrostatic interactions between the protein complex and the poly(3,4-ethylenedioxythiophene)-polystyrenesulfonate (PEDOT:PSS) polymer. Further, we follow a layer-by-layer (LBL) deposition procedure to produce up to 9-layer pairs of PSI and PEDOT:PSS with highly reproducible layer thicknesses as well as distinct changes in surface composition. When tested in an electrochemical cell employing ubiquinone-0 as a mediator, the photocurrent performance of the LBL films increased linearly by 83 ± 6 nA/cm2 per PSI layer up to 6-layer pairs. The 6-layer pair samples yielded a photocurrent of 414 ± 13 nA/cm2, after which the achieved photocurrent diminished with additional layer pairs. The turnover number (TN) of the PSI-PEDOT:PSS LBL assemblies also greatly exceeds that of drop-casted PSI multilayer films, highlighting that the rate of electron collection is improved through the systematic deposition of the protein complexes and conducting polymer. The capability to deposit high loadings of PSI between PEDOT:PSS layers, while retaining connection to the underlying electrode, shows the value in using LBL assembly to produce PSI and PEDOT:PSS bioelectrodes for photoelectrochemical energy harvesting applications.

Elevated levels of circulating cell-free hemoglobin (CFH) are an integral feature of several clinical conditions including sickle cell anemia, sepsis, hemodialysis and cardiopulmonary bypass. Oxidized (Fe

Here, we describe the surprising reactivity between surface-attached (a) 0.9, 1.6, and 4.1 nm diameter weakly stabilized Au nanoparticles (NPs) and aqueous 1.0 × 10-4 M Ag+ solution, and (b) 1.6 and 4.1 nm diameter weakly stabilized Au NPs and aqueous 1.0 × 10-5 M PtCl42-, which are considered to be antigalvanic replacement (AGR) reactions because they are not thermodynamically favorable for bulk-sized Au under these conditions. Anodic Stripping Voltammetry (ASV) and Scanning Transmission Electron Microscopy with Energy-Dispersive X-ray Spectroscopy (STEM-EDS) mapping provide quantitation of the extent of Ag and Pt replacement as a function of Au NP diameter. The extent of the reaction increases as the Au NP size decreases. The percentage of Ag in the AuAg alloy following AGR based on ASV is 17.8 ± 0.6% for 4.1 nm diameter Au NPs, 87.2 ± 2.9% for 1.6 nm Au NPs, and an unprecedented full 100% Ag for 0.9 nm diameter Au NPs. STEM-EDS mapping shows very close agreement with the ASV-determined compositions. In the case of PtCl42-, STEM-EDS mapping shows AuPt alloy NPs with 3.9 ± 1.3% and 41.1 ± 8.7% Pt following replacement with 4.1 and 1.6 nm diameter Au NPs, respectively, consistent with qualitative changes to the ASV. The size-dependent AGR correlates well with the negative shift in the standard potential (E0) for Au oxidation with decreasing NP size.

The photosystem I (PSI) protein complex is known to enhance bioelectrode performance for many liquid-based photoelectrochemical cells. A hydrogel as electrolyte media allows for simpler fabrication of more robust and practical solar cells in comparison to liquid-based devices. This paper reports a natural, gel-based dye-sensitized solar cell that integrates PSI to improve device efficiency. TiO

Isolated photosystem I (PSI) has been integrated into numerous technologies for solar energy conversion. Interest in PSI is a consequence of its high internal quantum efficiency, thermal stability, ease of extraction, and adaptability. While there has been success in improving performance to elevate PSI biohybrid technologies toward a practical realm, the stability of PSI bioelectrodes is also of critical importance. Commercial solar energy conversion technologies are expected to achieve lifetimes of the order of ten years; however, many research-scale PSI bioelectrodes have only been tested for tens of days. Key areas affecting PSI bioelectrode stability include the effects of reactive oxygen species, immobilization strategies, and the environment within solid-state PSI biohybrid photovoltaics. At the current state, further investigation of long-term stability is necessary in enabling the development of PSI bioelectrodes for both photoelectrochemical cells and solid-state biohybrid photovoltaics.

Carbon dots (CDs) are a rapidly progressing class of nanomaterial which show promise towards applications in solar energy conversion due to their low toxicity, favorable electrochemical properties, and tunability. In recent years there have been a number of reported CD syntheses, both top-down and bottom-up methods, producing a diverse range of CDs with intrinsic properties dependent on the starting materials and utilized dopants. This work presents a citrate buffer-facilitated synthesis of nitrogen-doped carbon dots (NCD) and explores the impact of urea concentration on observed electrochemical and optical properties. Optical absorbance and quantum yield of NCDs were found to increase with the dopant concentrations present in the hydrothermal reaction mixture. Electrochemical analysis demonstrates that increased nitrogen content results in the shifting of carbon dot oxidation potentials without the need of post-synthesis surface modifications. Over the range of molar ratios of dopant-to-citrate tested, the oxidation potentials of NCDs shifted up to 150 mV towards more negative potentials. X-ray photoelectron spectroscopy confirms the addition of pyrrolic and pyridinic nitrogen at different levels in different batches of NCDs, which are likely the source of the observed changes.

As energy consumption increases in tandem with the global warming crisis, the need for cleaner, renewable energy is at an all-time high. Inspired by nature’s billion-year-old process for energy production, biohybrid solar approaches have been shown to hold much promise in many aspects that are required in renewable energy. By leveraging Photosystem I (PSI), a key protein of the electron transport chain in photosynthesis, it is possible to hijack the electrons to perform an array of light-driven processes. In many areas of PSI research, conductive polymers are employed to wire the protein so that electrons can be directed to and from the reaction centers along favorable paths. Poly(3,4-ethylenedioxythiophene) polystyrene sulfonate (PEDOT:PSS) is commonly used in these applications as it is water soluble and thus compatible with PSI. Electro-polymerization allows for direct contact of the PEDOT with an electrode surface; however, electropolymerized PEDOT:PSS is mechanically unstable. Limited research has been conducted to demonstrate the effects of other counter anions on PEDOT properties and their implications on the incorporation of PSI into a hybrid device. In this work, we describe the methodologies employed to synthesize and characterize PEDOT thin films with varying counter anions. Our findings indicate that the counter anion during the electropolymerization process affects morphology, charge transfer kinetics, and optical properties of the resulting film. Coupling these observations to the photoactivity of PEDOT-PSI hybrid films, we provide explanations for the varied performance based upon fundamental understandings of chemical and electrochemical phenomena.

Preterm birth (PTB) is the top cause of death in newborns and the second leading cause of death in children under the age of five. Certain cytokines, like interleukin-6 (IL-6), and matrix metalloproteinases (MMPs) are known to facilitate the degradation of the fetal membrane, but the mechanisms that result in fetal membrane rupture under both normal and pathological conditions are largely unknown. Our fetal membrane-on-a-chip model will allow us to investigate the conditions and mechanisms that precede fetal membrane rupture. Our enzymatic electrochemical detection platform allows us to detect biosignatures from the fetal membrane by measuring analyte concentration changes such as glucose utilization, lactate production, and oxygen consumption. Addition of electrochemical sensors for cytokine and MMPs to the detection platform will provide further insight into the mechanisms underlying PTB by offering low-level protein biomarker quantification. We are currently fabricating and optimizing electrochemical immunoassays for the detection of IL-6 and MMP-3 to leverage the advantages of electrochemical techniques in studying preterm births. The current protein assay design utilizes the enzyme alkaline phosphatase attached to the target protein. When introducing the substrate p-aminophenyl phosphate (PAPP) to the system, it is enzymatically converted to p-aminophenol (PAP). Upon oxidation, PAP is converted to p-quinone (PQI). We are currently studying the electrochemical behavior of PAP/PQI for application in immunoassays. We are investigating the use of different electrochemical techniques for PAP oxidation to PQI including cyclic voltammetry, square wave voltammetry, differential pulse voltammetry, and normal pulse voltammetry to see how the technique affects the reproducibility of assay measurements.

Chlorpyrifos (CPF), an organophosphate pesticide widely used on food crops and in households, has been a critical public health concern due to its neurotoxic effects. Investigations of CPF-induced neurotoxicity have focused on dysregulation of acetylcholinergic signaling but recent work suggests that alterations in glutamatergic neurotransmission and astrocytic glutamate metabolism may occur. L-glutamate is a major excitatory neurotransmitter that is mostly removed from the synaptic cleft by astrocytic glutamate transporters. Here we assessed the effects of CPF on net glutamate uptake by human-induced pluripotent stem cell derived astrocytes using electrochemical analysis. Our microclinical analyzer (µCA) paired with screen-printed electrodes (SPE) offers an automated sensing system that can be used to monitor cellular metabolism and signaling. In this work, a platinum based SPE combined with the µCA was used to electrochemically quantify glutamate uptake in human astrocytes. Immobilized glutamate oxidase on electrode surfaces provided stable and sensitive glutamate detection. The temperature modification of these electrodes (37°C) led to a nearly four-fold improvement in sensitivity (0.84±0.04 nA/µM), with low limits of detection (2.8±0.1 µM), and quantitation (9.3±0.5 µM). The linear range of the sensor (1-667 µM) is on the same order as typical extracellular glutamate concentrations. These sensors were used to assess the net glutamate uptake efficiency by using escalating concentrations of glutamate (10-200 µM). Glutamate uptake was most efficient when astrocytes were exposed to 50 µM glutamate for 30min (50% net uptake). We are investigating acute and chronic effects of net glutamate uptake using sublethal doses of CPF (0-100 µM). Identifying these cellular mechanisms affected by CPF is critical for the understanding of the impact of organophosphate toxicity.

Photosystem I (PSI) is one of the primary macromolecular machines that drive photosynthesis in green plants and cyanobacteria. Extracted PSI has been employed successfully as a macromolecular photosensitizer within a host of low-cost electrochemical and solid-state photovoltaic architectures. This presentation will also explore our group’s recent efforts to integrate PSI with advanced nanomaterials, including carbon nanotubes, carbon quantum dots, and conducting polymers polyaniline (PANI), polypyrrole, polyviologens, and poly(3,4-ethylenedioxythiophene). These composite assemblies enhance charge shuttling processes from individual proteins within multilayer assemblies—greatly reducing charge transfer resistances and improving overall efficiency of photocells. The group has reported two new prototype solid-state devices in which PSI or PSI/PANI is sandwiched between energetically appropriate electrodes. The group has also succeeded in stabilizing PSI films via crosslinking to create “wet” photoelectrochemical cells with greater performance and longevity. Finally, our current work is aimed at building new prototypes using PSI in solid state interfaces for scalable solar energy conversion. Finally, the incorporation of PSI into conducting polymer frameworks holds promise for improved conductivity and orientational control in the photoactive layers in these devices.

There is a need for valves and pumps that operate at the microscale with precision and accuracy, are versatile in their application, and are easily fabricated. To that end, we developed a new rotary planar multiport valve to faithfully select solutions (contamination = 5.22 ± 0.06 ppb) and a rotary planar peristaltic pump to precisely control fluid delivery (flow rate = 2.4 ± 1.7 to 890 ± 77 μL/min). Both the valve and pump were implemented in a planar format amenable to single-layer soft lithographic fabrication. These planar microfluidics were actuated by a rotary motor controlled remotely by custom software. Together, these two devices constitute an innovative microformulator that was used to prepare precise, high-fidelity mixtures of up to five solutions (deviation from prescribed mixture = ±|0.02 ± 0.02| %). This system weighed less than a kilogram, occupied around 500 cm(3), and generated pressures of 255 ± 47 kPa. This microformulator was then combined with an electrochemical sensor creating a microclinical analyzer (μCA) for detecting glutamate in real time. Using the chamber of the μCA as an in-line bioreactor, we compared glutamate homeostasis in human astrocytes differentiated from human-induced pluripotent stem cells (hiPSCs) from a control subject (CC-3) and a Tuberous Sclerosis Complex (TSC) patient carrying a pathogenic TSC2 mutation. When challenged with glutamate, TSC astrocytes took up less glutamate than control cells. These data validate the analytical power of the μCA and the utility of the microformulator by leveraging it to assess disease-related alterations in cellular homeostasis.

An amperometric acetylcholine sensor was developed for use in a microfluidic system and characterized with chlorpyrifos and its metabolite, chlorpyrifos oxon. This enzymatic sensor was highly selective for acetylcholine, with a detection limit of 0.2 μM and sensitivity of 1.7 nA μM−1 from 1–150 μM. Though chlorpyrifos had no effect on sensor function, chlorpyrifos oxon significantly inhibited response across a range of concentrations (0.5–50 μM). Inhibition was reversed by 2-pyridine aldoxime methyl chloride. This platform can be used to both quantify acetylcholine in the presence of chlorpyrifos and as a biorecognition method for chlorpyrifos oxon.

Dysregulations in metabolic and physiological cellular processes are generally accompanied by changes in concentration profiles of multiple biomarkers. Measuring dynamic changes in concentrations of several affected biomarkers can enhance our comprehension of the underlying disease pathophysiology and/or mechanisms of cytotoxicity. Most analytical detection techniques require samples to be obtained at pre-defined times for offline analysis. However, the inadequate temporal resolution of these methods can lead to key biological events that occur at shorter timescales, being overlooked. Electrochemical biosensors are relatively simple, cost-effective detection techniques that have been used extensively for continuous analyte monitoring, and more recently, for sensing multiple analytes. We describe the design and functionality of a novel electrochemical detection platform, comprising a screen-printed Pt electrode array, a microfluidic chip, and an automated pump and valve system, capable of quantitatively detecting up to eight analytes. As a proof of concept, we show the simultaneous, sensitive, and selective amperometric detection of four essential biomarkers- glucose, lactate, glutamate, and acetylcholine, in real time. We also demonstrate the biological application of this platform by continuously monitoring exocytotic glutamate release from human induced pluripotent stem cell-derived neuronal cultures over time. Excessive accumulation of glutamate in the synaptic cleft leads to prolonged stimulation of glutamate receptors on neurons, triggering a cascade of neurotoxic events that ultimately results in loss of neuronal function and cell death. The results suggest that our multianalyte electrochemical sensor platform can be coupled with more complex biological systems such as in vitro organotypic models, for the accurate monitoring of biomarker changes during adverse events like acute neurotoxicity and microbial infection.

The photosynthetic protein complex, photosystem I (PSI), can be photoexcited with a quantum efficiency approaching unity and can be integrated into solar energy conversion devices as the photoactive electrode. The incorporation of PSI into conducting polymer frameworks allows for improved conductivity and orientational control in the photoactive layer. Polyviologens are a unique class of organic polycationic polymers that can rapidly accept electrons from a primary donor such as photoexcited PSI and subsequently can donate them to a secondary acceptor. Monomeric viologens, such as methyl viologen, have been widely used as diffusible mediators in wet PSI-based photoelectrochemical cells on the basis of their suitable redox potentials for accepting electrons. Polyviologens possess similar electronic properties to their monomers with the added advantage that they can shuttle electrons in the solid state. Depositing polyviologen directly onto a film of PSI protein results in significant photocurrent enhancement, which confirms its role as an electron-transport material. The polymer film not only improves the photocurrent by aiding the electron transfer but also helps preserve the protein film underneath. The composite polymer-PSI assembly enhances the charge-shuttling processes from individual protein molecules within the PSI multilayer, greatly reducing charge-transfer resistances. The resulting PSI-based solid-state platform demonstrates a much higher photocurrent than the corresponding photoelectrochemical cell built using a similar architecture.

Electrochemical sensors are used by millions of patients and health care providers every year, yet these measurements are hindered by compounds that also exhibit inherent redox activity. Acetaminophen (APAP) is one such interferent that falls into this extensive class. In this work, an osmium-based redox polymer was used for electrochemical detection in a sensor that was operated at a decreased voltage, allowing for decreased interference. These sensors demonstrated better selectivity (40-fold for glucose and 200-fold for lactate) for their respective analyte over APAP, possessed higher sensitivity (0.350 ± 0.006 μA mM-1 for glucose and 2.00 ± 0.05 μA mM-1 for lactate) over a broad range of analyte concentrations (50 μM-10 mM for glucose and 2-324 μM for lactate), and displayed similar operational stability (26% decrease for glucose and 29% decrease for lactate) over 7 days compared to first-generation sensors. To test this platform under biologically-relevant conditions, glucose metabolism was monitored in a model liver cell line, Alpha Mouse Liver 12 (AML12) after treatment with APAP and/or insulin. This work represents a high-resolution electrochemical biosensor for microphysiological monitoring of glucose and lactate in the presence of APAP.

Electrochemical sensors that utilize enzymes are a sensitive, inexpensive means of detecting biologically relevant analytes. These sensors are categorized based on their construction and method of signal transport. Type I sensors consist of a crosslinked enzyme on an electrode surface and are potentially subject to interference from byproducts and other biological analytes. However, type II sensors help alleviate this problem with the addition of a redox polymer layer that assists in signal transduction, thus minimizing interferences. An osmium-loaded poly(vinylimidazole) polymer (Os-PVI) is commonly used with successful results, and when combined with an enzyme yields a type II sensor. Our initial attempts at the synthesis of this polymer resulted in an unexpected osmium precursor, which had fluorescent and redox properties that did not match with the desired Os-PVI polymer. Careful exclusion of oxygen during the Os complex precursor synthesis was necessary to avoid this unexpected oxygen containing Os-precursor, which had been seen previously in mass spectrometry studies. All precursors and osmium polymers were characterized with 1H NMR, fluorescence, mass spectrometry, and cyclic voltammetry to provide a better understanding of these compounds and assist in the building of new sensors.

Preterm birth (PTB) is clinically defined as process of giving birth before 37 weeks of gestation and is a leading cause of death among neonates and children under the age of five. Prematurity remains a critical issue in developed countries, yet our understanding of the pathophysiology of PTB remains largely unknown. Among pregnancy complications, subclinical infections such as chorioamnionitis (CAM) are implicated in up to 70% of PTB cases. Specifically, CAM is characterized by the infection of the fetal membranes that surround the developing fetus and extend from the placenta, and is often associated with preterm, premature rupture of the fetal membranes (PPROM). The fetal membrane plays a key structural role in maintaining the fetal and maternal compartments of the gravid uterus. However, our understanding of the mechanisms of PPROM and the spatio-temporal progress of CAM remains vastly unknown. A lack of human-derived models have hindered our understanding of the mechanism that govern spontaneous PTB. Thus, in this short review, we discuss the emerging microfabrication technologies, specifically, organ-on-chip (OoCs) models, that seek to recapitulate the cellular and molecular context of the gestational membranes in vitro. These models show promise to facilitate the investigation of pathologic mechanisms that drive these disease conditions by mimicking the interactive contribution of the major cell types that make up the microenvironment of the fetal membrane and enable high throughput screening. Herein, we histologically characterize the microenvironment of the fetal membrane as a metric for scaling to recapitulate the functional components of the human fetal membrane. We review the current OoC models of the gravid uterus and conceptualize an "Instrumented Fetal Membrane on a Chip" (IFMOC) design as a prototype for PPROM and CAM research. Lastly, we discuss further applications of these OoC models for toxicological or pharmacological screening and personalized medicine. Fetal membrane OoCs offer an innovative and valuable platform to explore complex interactions between multiple drug types, toxic substances, and/or pathogenic microbes and their potential impacts on pregnancy outcomes. Further work will be required by integrating technological and analytical capabilities in order to characterize the fetal membrane microenvironment for preterm birth research.

Here, a new design for natural dye-sensitized solar cells is presented. A stacked assembly consisting of a discrete Photosystem I protein multilayer film atop a natural dye-sensitized photoanode expands absorbance of the solar spectrum to facilitate a more than 2-fold increase in cell photovoltage relative to the unmodified equivalent.

2020 H. H. Uhlig Summer Research Fellowship – Summary Report

Summary The abundant photosynthetic protein Photosystem I (PSI) achieves 1.1 V charge separation with nearly perfect quantum yield. Researchers from around the world have extracted PSI from photosynthetic organisms for use in biohybrid photoelectrochemical systems. To this end, PSI has been paired with a host of conductive matrices for progressively more effective energy conversion based on mediated approaches to protein–electrode charge exchange. In this review, we highlight the use of films containing PSI for direct solar energy conversion in electrochemical and solid-state arrangements. We emphasize recent advances in pairing PSI proteins with both ion- and electron-conducting polymer scaffolds.

Extracted from the chloroplasts of green plants, Photosystem I (PSI) is a multi-subunit protein complex that can be deposited as a thin film onto electrodes and incorporated into biohybrid photovoltaic devices to convert sunlight into usable energy. Previously, our group characterized persistent photocurrents over nine months using PSI multilayer films in an aqueous solution of redox mediators. However, the loss of periphery PSI proteins upon exposure to mediator solutions is known to affect the photocatalytic performance of the overall film. The cross-linker 2-iminothiolane (2IT) is capable of connecting protein molecules with covalent bonds, and is herein used to stabilize PSI multilayer films on gold substrates against significant desorption/degradation. Results showed that the thickness retainment of the cross-linked PSI films was superior to that of non-cross-linked films. PSI films modified by 2IT maintained production of photocurrent with long-term exposure to aqueous solutions in contrast to non-cross-linked control films.

Objective: Continuous glucose monitor (CGM) measured glucose lags blood glucose by several minutes, and this delay is usually attributed to the latency of interstitial fluid. However, independent findings suggest that interstitial glucose dispersion is near-immediate. This study seeks to determine the physiological locus of CGM latency. Methods: Current was measured at 1 Hz in Dexcom G4 CGMs using a CHI 1440 potentiostat. CGMs were exposed to varying glucose levels in vitro to define sensor kinetics. Next, CGMs were implanted in mice, and sensor vs. blood glucose responses were measured following an intravenous glucose challenge. Finally, dispersion of a fluorescent glucose analogue (2-NBDG) into the CGM micro-environment was observed in vivo using intravital microscopy. Tissue deposited on sensors and non-CGM implanted subcutaneous fat were then collected for histological analysis. Results: The CGM time constant in vitro was 51±3 seconds. In vivo, CGMs took 24±7 minutes to reach a local maximum in maximum current following intravenous injection of glucose. 2-NBDG took 21±7 minutes to reach maximum fluorescence near the sensor, vs. 6±6 minutes in subcutaneous fat (p=0.0011). Collagen was increased in sensor deposits relative to non-CGM implanted subcutaneous fat (p=0.0018), and collagen content was correlated with 2-NBDG latency (R=0.96, p=0.0004). Conclusions: Interstitial glucose responds to changes in blood glucose within seconds, whereas the glucose levels seen by a CGM are delayed by several minutes due to fibrous encapsulation. A CGM that avoids encapsulation could approximate real-time blood glucose measurements. Additionally, these data raise the possibility that fibrosis underlies insulin resistance by creating a biophysical impediment to the interstitial dispersion of glucose. Disclosure P.M. McClatchey: None. E.S. McClain: None. I.M. Williams: None. J.M. Gregory: Consultant; Self; InClinica. D. Cliffel: None. D. Wasserman: None. Funding National Institutes of Health (DK059637, DK054902, DK050277, T32DK101003, F32DK120104); American Heart Association

Continuous glucose monitor (CGM) readings are delayed relative to blood glucose, and this delay is usually attributed to the latency of interstitial glucose levels. However, CGM-independent data suggest rapid equilibration of interstitial glucose. This study sought to determine the loci of CGM delays. Electrical current was measured directly from CGM electrodes to define sensor kinetics in the absence of smoothing algorithms. CGMs were implanted in mice, and sensor versus blood glucose responses were measured after an intravenous glucose challenge. Dispersion of a fluorescent glucose analog (2-NBDG) into the CGM microenvironment was observed in vivo using intravital microscopy. Tissue deposited on the sensor and nonimplanted subcutaneous adipose tissue was then collected for histological analysis. The time to half-maximum CGM response in vitro was 35 ± 2 s. In vivo, CGMs took 24 ± 7 min to reach maximum current versus 2 ± 1 min to maximum blood glucose (P = 0.0017). 2-NBDG took 21 ± 7 min to reach maximum fluorescence at the sensor versus 6 ± 6 min in adipose tissue (P = 0.0011). Collagen content was closely correlated with 2-NBDG latency (R = 0.96, P = 0.0004). Diffusion of glucose into the tissue deposited on a CGM is substantially delayed relative to interstitial fluid. A CGM that resists fibrous encapsulation would better approximate real-time deviations in blood glucose.

The ability to assess oxygenation within living cells is much sought after to more deeply understand normal and pathological cell biology. Hypoxia Red manufactured by Enzo Life Sciences is advertised as a novel hypoxia detector dependent on nitroreducatase activity. We sought to use Hypoxia Red in primary neuronal cultures to test cell-to-cell metabolic variability in response to hypoxic stress. Neurons treated with 90 min of hypoxia were labeled with Hypoxia Red. We observed that, even under normoxic conditions neurons expressed fluorescence robustly. Analysis of the chemical reactions and biological underpinnings of this method revealed that the high uptake and reduction of the dye is due to active nitroreductases in normoxic cells that are independent of oxygen availability.

Globally, maternal and fetal health is greatly impacted by extraplacental inflammation. Group B Streptococcus (GBS), a leading cause of chorioamnionitis, is thought to take advantage of the uterine environment during pregnancy in order to cause inflammation and infection. In this study, we demonstrate the metabolic changes of murine macrophages caused by GBS exposure. GBS alone prompted a delayed increase in lactate production, highlighting its ability to redirect macrophage metabolism from aerobic to anaerobic respiration. This production of lactate is thought to aid in the development and propagation of GBS throughout the surrounding tissue. Additionally, this study shows that PGE2 priming was able to exacerbate lactate production, shown by the rapid and substantial lactate increases seen upon GBS exposure. These data provide a novel model to study the role of GBS exposure to macrophages with and without PGE2 priming.

We have built a multianalyte microphysiometer that detects multiple analytes involved in the cellular bioenergetics simultaneously. Monitoring the uptake of glucose and oxygen, and the production of lactate and acid gives a more complete picture of the metabolic processes within the cell than just acidification microphysiometry alone. Metabolic processes such as glycolysis, mitochondrial ATP generation, and glycogenesis are all directly related to the flux of these analytes. Temporal resolution of metabolic responses is much faster than conventional well-plate studies, leading to dynamic metabolic data. Electrochemical sensors are easily adapted into microfluidic formats for microphysiology readouts. Our newest instrumental designs include a complete overhaul of the microfluidic chambers, screen printed electrodes, and microfluidic pumps, valves, and drivers that will enable easy adoption of this methodology in the development of 3D cell and tissue studies using 8 separate electrochemical measurements at a time. In the first design, 8 separate amperometric sensors are developed using a screen printed electrode with isolated microfluidic channels. In the second design, 4 amperometric sensors are combined with 4 potentiometric sensors using the newly created CH1440 created by CH Instruments. The adaption of this technology to instrumenting organs-on-a-chip is currently underway.

Biohybrid photovoltaics leverage naturally occurring photo-active protein complexes to convert solar energy into electricity; offering a unique route to lowering the cost and environmental impact of solar energy conversion. Photosystem I (PSI), a protein complex found in higher plants, algae, and cyanobacteria, produces photoexcited excitons over a distance of approximately 10 nm with a 1.1 V photopotential. PSI has been utilized in a variety of biohybrid photovoltaics with success; however, improvements in performance are necessary to bring PSI – based devices to a technologically viable state. The electrons and holes produced within a PSI biohybrid photovoltaic must pass through multiple interfaces before being harnessed in an electrical circuit. Therefore, the total current, potential, and efficiency are dependent on losses incurred at each interface and could be vastly improved by optimizing individual electron transfer steps. Many researchers are looking toward direct electron transfer (DET) between the active sites of PSI and electrodes to reduce these losses. Unfortunately, achieving DET is difficult due to the necessity of proper PSI orientation and the close proximity (