Space radiation damage rescued by inhibition of key spaceflight associated miRNAs.

Our previous research revealed a key microRNA signature that is associated with spaceflight that can be used as a biomarker and to develop countermeasure treatments to mitigate the damage caused by space radiation. Here, we expand on this work to determine the biological factors rescued by the countermeasure treatment. We performed RNA-sequencing and transcriptomic analysis on 3D microvessel cell cultures exposed to simulated deep space radiation (0.5 Gy of Galactic Cosmic Radiation) with and without the antagonists to three microRNAs: miR-16-5p, miR-125b-5p, and let-7a-5p (i.e., antagomirs). Significant reduction of inflammation and DNA double strand breaks (DSBs) activity and rescue of mitochondria functions are observed after antagomir treatment. Using data from astronaut participants in the NASA Twin Study, Inspiration4, and JAXA missions, we reveal the genes and pathways implicated in the action of these antagomirs are altered in humans. Our findings indicate a countermeasure strategy that can potentially be utilized by astronauts in spaceflight missions to mitigate space radiation damage.

Aging and putative frailty biomarkers are altered by spaceflight.

Human space exploration poses inherent risks to astronauts' health, leading to molecular changes that can significantly impact their well-being. These alterations encompass genomic instability, mitochondrial dysfunction, increased inflammation, homeostatic dysregulation, and various epigenomic changes. Remarkably, these changes bear similarities to those observed during the aging process on Earth. However, our understanding of the connection between these molecular shifts and disease development in space remains limited. Frailty syndrome, a clinical syndrome associated with biological aging, has not been comprehensively investigated during spaceflight. To bridge this knowledge gap, we leveraged murine data obtained from NASA's GeneLab, along with astronaut data gathered from the JAXA and Inspiration4 missions. Our objective was to assess the presence of biological markers and pathways related to frailty, aging, and sarcopenia within the spaceflight context. Through our analysis, we identified notable changes in gene expression patterns that may be indicative of the development of a frailty-like condition during space missions. These findings suggest that the parallels between spaceflight and the aging process may extend to encompass frailty as well. Consequently, further investigations exploring the utility of a frailty index in monitoring astronaut health appear to be warranted.

The interplay between lncRNAs, RNA-binding proteins and viral genome during SARS-CoV-2 infection reveals strong connections with regulatory events involved in RNA metabolism and immune response.

Rationale: Viral infections are complex processes based on an intricate network of molecular interactions. The infectious agent hijacks components of the cellular machinery for its profit, circumventing the natural defense mechanisms triggered by the infected cell. The successful completion of the replicative viral cycle within a cell depends on the function of viral components versus the cellular defenses. Non-coding RNAs (ncRNAs) are important cellular modulators, either promoting or preventing the progression of viral infections. Among these ncRNAs, the long non-coding RNA (lncRNA) family is especially relevant due to their intrinsic functional properties and ubiquitous biological roles. Specific lncRNAs have been recently characterized as modulators of the cellular response during infection of human host cells by single stranded RNA viruses. However, the role of host lncRNAs in the infection by human RNA coronaviruses such as SARS-CoV-2 remains uncharacterized. Methods: In the present work, we have performed a transcriptomic study of a cohort of patients with different SARS-CoV-2 viral load and analyzed the involvement of lncRNAs in supporting regulatory networks based on their interaction with RNA-binding proteins (RBPs). Results: Our results revealed the existence of a SARS-CoV-2 infection-dependent pattern of transcriptional up-regulation in which specific lncRNAs are an integral component. To determine the role of these lncRNAs, we performed a functional correlation analysis complemented with the study of the validated interactions between lncRNAs and RBPs. This combination of in silico functional association studies and experimental evidence allowed us to identify a lncRNA signature composed of six elements - NRIR, BISPR, MIR155HG, FMR1-IT1, USP30-AS1, and U62317.2 - associated with the regulation of SARS-CoV-2 infection. Conclusions: We propose a competition mechanism between the viral RNA genome and the regulatory lncRNAs in the sequestering of specific RBPs that modulates the interferon response and the regulation of RNA surveillance by nonsense-mediated decay (NMD).

Measuring Spatial and Temporal Oxygen Flux Near Plant Tissues Using a Self-Referencing Optrode.

Self-referencing optrodic microsensing is a noninvasive method for measuring oxygen transport into/from tissues. The sensing mechanism is based on fluorescence quenching by molecular oxygen at the tip of a fiber-optic probe, and facilitates microscale spatial mapping and continuous monitoring at 100-350 mHz sampling frequency. Over the last decade, this technique has been applied for plant tissues, including roots, seeds, leaves, and flowers in both liquid and air. Here, we describe the operating principle of self-referencing optrodic microsensing for the study of plant tissues with a specific focus on juvenile roots.

An assessment of US microbiome research.

Genome-enabled technologies have supported a dramatic increase in our ability to study microbial communities in environments and hosts. Taking stock of previously funded microbiome research can help to identify common themes, under-represented areas and research priorities to consider moving forward. To assess the status of US microbiome research, a team of government scientists conducted an analysis of federally funded microbiome research. Microbiomes were defined as host-, ecosystem- or habitat-associated communities of microorganisms, and microbiome research was defined as those studies that emphasize community-level analyses using 'omics technologies. Single pathogen, single strain and culture-based studies were not included, except symbiosis studies that served as models for more complex communities. Fourteen governmental organizations participated in the data call. The analysis examined three broad research themes, eight environments and eight microbial categories. Human microbiome research was larger than any other environment studied, and the basic biology research theme accounted for half of the total research activities. Computational biology and bioinformatics, reference databases and biorepositories, standardized protocols and high-throughput tools were commonly identified needs. Longitudinal and functional studies and interdisciplinary research were also identified as needs. This study has implications for the funding of future microbiome research, not only in the United States but beyond.

Sodium channel Nax is a regulator in epithelial sodium homeostasis.

The mechanisms by which the epidermis responds to disturbances in barrier function and restores homeostasis are unknown. With a perturbation of the epidermal barrier, water is lost, resulting in an increase in extracellular sodium concentration. We demonstrate that the sodium channel Nax functions as a sodium sensor. With increased extracellular sodium, Nax up-regulates prostasin, which results in activation of the sodium channel ENaC, resulting in increased sodium flux and increased downstream mRNA synthesis of inflammatory mediators. Nax is present in multiple epithelial tissues, and up-regulation of its downstream genes is found in hypertrophic scars. In animal models, blocking Nax expression results in improvement in scarring and atopic dermatitis-like symptoms, both of which are pathological conditions characterized by perturbations in barrier function. These findings support an important role for Nax in maintaining epithelial homeostasis.

Altered glucose metabolism in Harvey-ras transformed MCF10A cells.

Metabolic reprogramming that alters the utilization of glucose including the "Warburg effect" is critical in the development of a tumorigenic phenotype. However, the effects of the Harvey-ras (H-ras) oncogene on cellular energy metabolism during mammary carcinogenesis are not known. The purpose of this study was to determine the effect of H-ras transformation on glucose metabolism using the untransformed MCF10A and H-ras oncogene transfected (MCF10A-ras) human breast epithelial cells, a model for early breast cancer progression. We measured the metabolite fluxes at the cell membrane by a selective micro-biosensor, [(13)C6 ]glucose flux by (13)C-mass isotopomer distribution analysis of media metabolites, intracellular metabolite levels by NMR, and gene expression of glucose metabolism enzymes by quantitative PCR. Results from these studies indicated that MCF10A-ras cells exhibited enhanced glycolytic activity and lactate production, decreased glucose flux through the tricarboxylic acid (TCA) cycle, as well as an increase in the utilization of glucose in the pentose phosphate pathway (PPP). These results provide evidence for a role of H-ras oncogene in the metabolic reprogramming of MCF10A cells during early mammary carcinogenesis.

Glutathione-gated potassium efflux as a mechanism of active biofilm detachment.

Biofilm detachment often has detrimental effects such as pipe obstruction and infection, yet the detachment mechanisms underlying dispersal remain largely unknown. In this study, a stress response mechanism known as glutathione-gated potassium efflux (GGKE) was evaluated as an active detachment mechanism in the dispersal of Pseudomonas aeruginosa biofilms. N-ethylmaleimide (NEM) was used to activate potassium efflux proteins (Kef) associated with the GGKE pathway. This stress response mechanism was hypothesized to lead to altered cation concentration, which can potentially affect polymer bridging in biofilms, and ultimately cause biofilm detachment. Results showed the activation of GGKE by NEM exposure caused biofilm detachment without inducing a measurable change in viability, and detached biomass concentration and composition were dependent on NEM concentration. More detached biomass was observed with higher concentrations of NEM, with a trend of increasing polymer detachment. The detachment was likely resulting from a weakened biofilm structural integrity induced by bridge denaturing from GGKE activation. This study is important in understanding biofilm detachment from engineered systems such as membrane aerated bioreactors.

Comparative study of non-invasive methods for assessing Daphnia magna embryo toxicity.

Embryos, unlike adults, are typically sessile, which allows for an increase in the available metrics that can be used to assess chemical toxicity. We investigate Daphnia magna development rate and oxygen consumption as toxicity metrics and compare them to arrested embryo development using four different techniques with potassium cyanide (KCN) as a common toxicant. The EC50 (95 % CI) for arrested development was 2,535 (1,747-3,677) µg/L KCN. Using pixel intensity changes, recorded with difference imaging, we semi-quantitatively assessed a decrease in development rate at 200 µg/L KCN, threefold lower than the arrested development lowest observed effect concentration (LOEC). Respirometry and self-referencing (SR) microsensors were two unique techniques used to assess oxygen consumption. Using respirometry, an increase in oxygen consumption was found in the 5 µg/L KCN treatment and a decrease for 148 µg/L, but no change was found for the 78 µg/L KCN treatment. Whereas, with SR microsensors, we were able to detect significant changes in oxygen consumption for all three treatments: 5, 78, and 148 µg/L KCN. While SR offered the highest sensitivity, the respirometry platform developed for this study was much easier to use to measure the same endpoint. Oxygen consumption may be subject to change during the development process, meaning consumption assessment techniques may only be useful only for short-term experiments. Development rate was a more sensitive endpoint though was only reliable four of the six embryonic developmental stages examined. Despite being the least sensitive endpoint, arrested embryo development was the only technique capable of assessing the embryos throughout all developmental stages. In conclusion, each metric has advantages and limitations, but because all are non-invasive, it is possible to use any combination of the three.

Single-stranded DNA functionalized single-walled carbon nanotubes for microbiosensors via layer-by-layer electrostatic self-assembly.

In this letter, the facial noncovalent adsorption of single-stranded DNA (ssDNA) provided single-walled carbon nanotubes (SWNTs) with biofunctionality while their superior properties were retained. In this case, we innovatively demonstrated the feasibility of employing the negative surface charge of ssDNA-SWNTs to realize layer-by-layer electrostatic self-assembly. On the basis of such a sandwichlike structure, an applicable glucose microbiosensor with direct electrochemistry and high performance was fabricated. The proposed protocol provided an ideal platform for various sensing applications, and might have profound influence on related nanotechnology.

Silver nanoparticle-specific mitotoxicity in Daphnia magna.

Silver nanoparticles (Ag NPs) are gaining popularity as bactericidal agents in commercial products; however, the mechanisms of toxicity (MOT) of Ag NPs to other organisms are not fully understood. It is the goal of this research to determine differences in MOT induced by ionic Ag(+) and Ag NPs in Daphnia magna, by incorporating a battery of traditional and novel methods. Daphnia embryos were exposed to sublethal concentrations of AgNO3 and Ag NPs (130-650 ng/L), with uptake of the latter confirmed by confocal reflectance microscopy. Mitochondrial function was non-invasively monitored by measuring proton flux using self-referencing microsensors. Proton flux measurements revealed that while both forms of silver significantly affected proton efflux, the change induced by Ag NPs was greater than that of Ag(+). This could be correlated with the effects of Ag NPs on mitochondrial dysfunction, as determined by confocal fluorescence microscopy and JC-1, an indicator of mitochondrial permeability. However, Ag(+) was more efficient than Ag NPs at displacing Na(+) within embryonic Daphnia, based on inductively coupled plasma-mass spectroscopy (ICP-MS) analysis. The abnormalities in mitochondrial activity for Ag NP-exposed organisms suggest a nanoparticle-specific MOT, distinct from that induced by Ag ions. We propose that the MOT of each form of silver are complementary, and can act in synergy to produce a greater toxic response overall.

Mouse and human islets survive and function after coating by biosilicification.

Inorganic materials have properties that can be advantageous in bioencapsulation for cell transplantation. Our aim was to engineer a hybrid inorganic/soft tissue construct by inducing pancreatic islets to grow an inorganic shell. We created pancreatic islets surrounded by porous silica, which has potential application in the immunoprotection of islets in transplantation therapies for type 1 diabetes. The new method takes advantage of the islet capsule surface as a template for silica formation. Mouse and human islets were exposed to medium containing saturating silicic acid levels for 9-15 min. The resulting tissue constructs were then cultured for up to 4 wk under normal conditions. Scanning electron microscopy and energy dispersive X-ray spectroscopy was used to monitor the morphology and elemental composition of the material at the islet surface. A cytokine assay was used to assess biocompatibility with macrophages. Islet survival and function were assessed by confocal microscopy, glucose-stimulated insulin release assays, oxygen flux at the islet surface, expression of key genes by RT-PCR, and syngeneic transplant into diabetic mice.

Multi-analyte biochip (MAB) based on all-solid-state ion-selective electrodes (ASSISE) for physiological research.

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.

1,25-dihydroxyvitamin D regulation of glucose metabolism in Harvey-ras transformed MCF10A human breast epithelial cells.

This study was designed to investigate the impact of 1,25-dihydroxyvitamin D (1,25(OH)2D) on glucose metabolism during early cancer progression. Untransformed and ras-oncogene transfected (ras) MCF10A human breast epithelial cells were employed to model early breast cancer progression. 1,25(OH)2D modified the response of the ras cells to glucose restriction, suggesting 1,25(OH)2D may reduce the ras cell glucose addiction noted in cancer cells. To understand the 1,25(OH)2D regulation of glucose metabolism, following four-day 1,25(OH)2D treatment, metabolite fluxes at the cell membrane were measured by a nanoprobe biosensor, [(13)C6]glucose flux by (13)C-mass isotopomer distribution analysis of media metabolites, intracellular metabolite levels by NMR, and gene expression of related enzymes was assessed. Treatment with 1,25(OH)2D reduced glycolysis as flux of glucose to 3-phosphoglycerate was reduced by 15% (P=0.017) and 32% (P<0.003) in MCF10A and ras cells respectively. In the ras cells, 1,25(OH)2D reduced lactate dehydrogenase activity by 15% (P<0.05) with a concomitant 10% reduction in the flux of glucose to lactate (P=0.006), and reduction in the level of intracellular lactate by 55% (P=0.029). Treatment with 1,25(OH)2D reduced flux of glucose to acetyl-coA 24% (P=0.002) and 41% (P<0.001), and flux to oxaloacetate 33% (P=0.003) and 34% (P=0.027) in the MCF10A and ras cells, respectively, suggesting a reduction in tricarboxylic acid (TCA) cycle activity. The results suggest a novel mechanism involving the regulation of glucose metabolism by which 1,25(OH)2D may prevent breast cancer progression.

RNA-seq analysis identifies potential modulators of gravity response in spores of Ceratopteris (Parkeriaceae): evidence for modulation by calcium pumps and apyrase activity.

PREMISE OF THE STUDY: Gravity regulates the magnitude and direction of a trans-cell calcium current in germinating spores of Ceratopteris richardii. Blocking this current with nifedipine blocks the spore's downward polarity alignment, a polarization that is fixed by gravity ∼10 h after light induces the spores to germinate. RNA-seq analysis at 10 h was used to identify genes potentially important for the gravity response. The data set will be valuable for other developmental and phylogenetic studies. METHODS: De novo Newbler assembly of 958 527 reads from Roche 454 sequencing was executed. The sequences were identified and analyzed using in silico methods. The roles of endomembrane Ca(2+)-ATPase pumps and apyrases in the gravity response were further tested using pharmacological agents. KEY RESULTS: Transcripts related to calcium signaling and ethylene biosynthesis were identified as notable constituents of the transcriptome. Inhibiting the activity of endomembrane Ca(2+)-ATPase pumps with 2,5-di-(t-butyl)-1,4-hydroquinone diminished the trans-cell current, but increased the orientation of the polar axis to gravity. The effects of applied nucleotides and purinoceptor antagonists gave novel evidence implicating extracellular nucleotides as regulators of the gravity response in these fern spores. CONCLUSIONS: In addition to revealing general features of the transcriptome of germinating spores, the results highlight a number of calcium-responsive and light-receptive transcripts. Pharmacologic assays indicate endomembrane Ca(2+)-ATPases and extracellular nucleotides may play regulatory roles in the gravity response of Ceratopteris spores.

Non-invasive flux measurements using microsensors: theory, limitations, and systems.

Knowledge of the fluxes of ions and neutral molecules across the outer membrane or boundary of living tissues and cells is an important strand of applied molecular biology. Such fluxes can be measured non-invasively with good resolution in time and space. Two systems (MIFE™ and SIET) have been developed and have become widely used to implement this technique, and they are commercially available. This Chapter is the first comparative description of these two systems. It gives the context, the basic underlying theory, practical limitations inherent in the technique, theoretical developments, guidance on the practicalities of the technique, and the functionality of the two systems. Although the technique is strongly relevant to plant salt tolerance and other plant stresses (drought, temperature, pollutants, waterlogging), it also has rich relevance throughout biomedical studies and the molecular genetics of transport proteins.

Abnormal mitochondrial function impairs calcium influx in diabetic mouse pancreatic beta cells.

BACKGROUND: Abnormal insulin secretion of pancreatic beta cells is now regarded as the more primary defect than the insulin function in the etiology of type 2 diabetes. Previous studies found impaired mitochondrial function and impaired Ca(2+) influx in beta cells in diabetic patients and animal models, suggesting a role for these processes in proper insulin secretion. The aim of this study was to investigate the detailed relationship of mitochondrial function, Ca(2+) influx, and defective insulin secretion. METHODS: We investigated mitochondrial function and morphology in pancreatic beta cell of diabetic KK-Ay mice and C57BL/6J mice. Two types of Ca(2+) channel activities, L-type and store-operated Ca(2+) (SOC), were evaluated using whole-cell patch-clamp recording. The glucose induced Ca(2+) influx was measured by a non-invasive micro-test technique (NMT). RESULTS: Mitochondria in KK-Ay mice pancreatic beta cells were swollen with disordered cristae, and mitochondrial function decreased compared with C57BL/6J mice. Ca(2+) channel activity was increased and glucose induced Ca(2+) influx was impaired, but could be recovered by genipin. CONCLUSION: Defective mitochondrial function in diabetic mice pancreatic beta cells is a key cause of abnormal insulin secretion by altering Ca(2+) influx, but not via Ca(2+) channel activity.

Mechanical stretch induced calcium efflux from bone matrix stimulates osteoblasts.

The mechanisms by which bone cells sense critically loaded regions of bone are still a matter of ongoing debate. Animal models to investigate response to microdamage involve post mortem immunohistological analysis and do not allow real-time monitoring of cellular response during the emergence of the damage in bone. Most in vitro mechanical stimulation studies are conducted on non-bone substrates, neglecting the damage-related alterations in the pericellular niche and their potential effects on bone cells. The current study reports spontaneous efflux of calcium ions (Ca(2+)) (1.924±0.742 pmol cm(-2)s(-1)) from regions of devitalized bone matrix undergoing post-yield strains, induced by a stress concentrator. When these samples are seeded with MC3T3-E1 osteoblasts, the strain-induced Ca(2+) efflux from bone elicits cell response at the stress concentration site as manifested by activation of intracellular calcium signaling (increase in fluorescence by 52%±27%). This activity is associated with extracellular calcium because the intracellular calcium signaling in response to mechanical loading subsides when experiments are repeated using demineralized bone substrates (increase in fluorescence by 6%±10%). These results imply a novel perspective where bone matrix acts as an intermediary mechanochemical transducer by converting mechanical strain into a chemical signal (pericellular calcium) to which cells respond. Such a mechanism may be responsible for triggering repair at locations of bone matrix undergoing critical deformation levels.

A comparative study of enzyme immobilization strategies for multi-walled carbon nanotube glucose biosensors.

This work addresses the comparison of different strategies for improving biosensor performance using nanomaterials. Glucose biosensors based on commonly applied enzyme immobilization approaches, including sol-gel encapsulation approaches and glutaraldehyde cross-linking strategies, were studied in the presence and absence of multi-walled carbon nanotubes (MWNTs). Although direct comparison of design parameters such as linear range and sensitivity is intuitive, this comparison alone is not an accurate indicator of biosensor efficacy, due to the wide range of electrodes and nanomaterials available for use in current biosensor designs. We proposed a comparative protocol which considers both the active area available for transduction following nanomaterial deposition and the sensitivity. Based on the protocol, when no nanomaterials were involved, TEOS/GOx biosensors exhibited the highest efficacy, followed by BSA/GA/GOx and TMOS/GOx biosensors. A novel biosensor containing carboxylated MWNTs modified with glucose oxidase and an overlying TMOS layer demonstrated optimum efficacy in terms of enhanced current density (18.3 ± 0.5 µA mM(-1) cm(-2)), linear range (0.0037-12 mM), detection limit (3.7 µM), coefficient of variation (2%), response time (less than 8 s), and stability/selectivity/reproducibility. H(2)O(2) response tests demonstrated that the most possible reason for the performance enhancement was an increased enzyme loading. This design is an excellent platform for versatile biosensing applications.

Microbiosensors based on DNA modified single-walled carbon nanotube and Pt black nanocomposites.

Glucose and ATP biosensors have important applications in diagnostics and research. Biosensors based on conventional materials suffer from low sensitivity and low spatial resolution. Our previous work has shown that combining single-walled carbon nanotubes (SWCNTs) with Pt nanoparticles can significantly enhance the performance of electrochemical biosensors. The immobilization of SWCNTs on biosensors remains challenging due to the aqueous insolubility originating from van der Waals forces. In this study, we used single-stranded DNA (ssDNA) to modify SWCNTs to increase solubility in water. This allowed us to explore new schemes of combining ssDNA-SWCNT and Pt black in aqueous media systems. The result is a nanocomposite with enhanced biosensor performance. The surface morphology, electroactive surface area, and electrocatalytic performance of different fabrication protocols were studied and compared. The ssDNA-SWCNT/Pt black nanocomposite constructed by a layered scheme proved most effective in terms of biosensor activity. The key feature of this protocol is the exploitation of ssDNA-SWCNTs as molecular templates for Pt black electrodeposition. The glucose and ATP microbiosensors fabricated on this platform exhibited high sensitivity (817.3 nA/mM and 45.6 nA/mM, respectively), wide linear range (up to 7 mM and 510 µM), low limit of detection (1 µM and 2 µM) and desirable selectivity. This work is significant to biosensor development because this is the first demonstration of ssDNA-SWCNT/Pt black nanocomposite as a platform for constructing both single-enzyme and multi-enzyme biosensors for physiological applications.