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Identification of phosphoproteins coupled to initiation of motility in live epididymal mouse sperm

A method for collecting live immotile cauda epididymal mouse sperm that initiate motility by dilution into an activation buffer is described. Sperm in collection buffer showed low percent motility (MOT) and population progression (PRG) that increased 10-fold and 9-fold, respectively, during the first 2 min after dilution into activation buffer. Western phosphoserine (pS), phosphothreonine (pT), and phosphotyrosine (pY) analysis revealed a 120 kDa protein that markedly increased in pT content during initiation of motility and may be related to FP130, the motility-coupled axonemal protein of sea urchin sperm. A prominent 82 kDa protein that was pS and pT-phosphorylated in immotile and motile sperm is likely the fibrous sheath component AKAP82 that is phosphorylated during spermatogenesis. Analysis of live human sperm also identified a prominent 120 kDa pT protein. Thus it appears that phosphorylation of FP130 and related 120 kDa proteins in mouse, and perhaps human sperm, represent common targets during motility initiation in sperm. Copyright 1998 Academic Press.

Non-NASA Center↗

Localized infusion of IGF-I results in skeletal muscle hypertrophy in rats

Insulin-like growth factor I (IGF-I) peptide levels have been shown to increase in overloaded skeletal muscles (G. R. Adams and F. Haddad. J. Appl. Physiol. 81: 2509-2516, 1996). In that study, the increase in IGF-I was found to precede measurable increases in muscle protein and was correlated with an increase in muscle DNA content. The present study was undertaken to test the hypothesis that direct IGF-I infusion would result in an increase in muscle DNA as well as in various measurements of muscle size. Either 0.9% saline or nonsystemic doses of IGF-I were infused directly into a non-weight-bearing muscle of rats, the tibialis anterior (TA), via a fenestrated catheter attached to a subcutaneous miniosmotic pump. Saline infusion had no effect on the mass, protein content, or DNA content of TA muscles. Local IGF-I infusion had no effect on body or heart weight. The absolute weight of the infused TA muscles was approximately 9% greater (P < 0.05) than that of the contralateral TA muscles. IGF-I infusion resulted in significant increases in the total protein and DNA content of TA muscles (P < 0.05). As a result of these coordinated changes, the DNA-to-protein ratio of the hypertrophied TA was similar to that of the contralateral muscles. These results suggest that IGF-I may be acting to directly stimulate processes such as protein synthesis and satellite cell proliferation, which result in skeletal muscle hypertrophy.

Non-NASA Center↗

Plant responses to environmental stress: regulation and functions of the Arabidopsis TCH genes

Expression of the Arabidopsis TCH genes is markedly upregulated in response to a variety of environmental stimuli including the seemingly innocuous stimulus of touch. Understanding the mechanism(s) and factors that control TCH gene regulation will shed light on the signaling pathways that enable plants to respond to environmental conditions. The TCH proteins include calmodulin, calmodulin-related proteins and a xyloglucan endotransglycosylase. Expression analyses and localization of protein accumulation indicates that the potential sites of TCH protein function include expanding cells and tissues under mechanical strain. We hypothesize that at least a subset of the TCH proteins may collaborate in cell wall biogenesis.

Review, Tutorial↗

Visualization of newt aragonitic otoconial matrices using transmission electron microscopy

Otoconia are calcified protein matrices within the gravity-sensing organs of the vertebrate vestibular system. These protein matrices are thought to originate from the supporting or hair cells in the macula during development. Previous studies of mammalian calcitic, barrel-shaped otoconia revealed an organized protein matrix consisting of a thin peripheral layer, a well-defined organic core and a flocculent matrix inbetween. No studies have reported the microscopic organization of the aragonitic otoconial matrix, despite its protein characterization. Pote et al. (1993b) used densitometric methods and inferred that prismatic (aragonitic) otoconia have a peripheral protein distribution, compared to that described for the barrel-shaped, calcitic otoconia of birds, mammals, and the amphibian utricle. By using tannic acid as a negative stain, we observed three kinds of organic matrices in preparations of fixed, decalcified saccular otoconia from the adult newt: (1) fusiform shapes with a homogenous electron-dense matrix; (2) singular and multiple strands of matrix; and (3) more significantly, prismatic shapes outlined by a peripheral organic matrix. These prismatic shapes remain following removal of the gelatinous matrix, revealing an internal array of organic matter. We conclude that prismatic otoconia have a largely peripheral otoconial matrix, as inferred by densitometry.

NASA Discipline Neuroscience↗

Structural characterization and regulatory element analysis of the heart isoform of cytochrome c oxidase VIa

In order to investigate the mechanism(s) governing the striated muscle-specific expression of cytochrome c oxidase VIaH we have characterized the murine gene and analyzed its transcriptional regulatory elements in skeletal myogenic cell lines. The gene is single copy, spans 689 base pairs (bp), and is comprised of three exons. The 5'-ends of transcripts from the gene are heterogeneous, but the most abundant transcript includes a 5'-untranslated region of 30 nucleotides. When fused to the luciferase reporter gene, the 3.5-kilobase 5'-flanking region of the gene directed the expression of the heterologous protein selectively in differentiated Sol8 cells and transgenic mice, recapitulating the pattern of expression of the endogenous gene. Deletion analysis identified a 300-bp fragment sufficient to direct the myotube-specific expression of luciferase in Sol8 cells. The region lacks an apparent TATA element, and sequence motifs predicted to bind NRF-1, NRF-2, ox-box, or PPAR factors known to regulate other nuclear genes encoding mitochondrial proteins are not evident. Mutational analysis, however, identified two cis-elements necessary for the high level expression of the reporter protein: a MEF2 consensus element at -90 to -81 bp and an E-box element at -147 to -142 bp. Additional E-box motifs at closely located positions were mutated without loss of transcriptional activity. The dependence of transcriptional activation of cytochrome c oxidase VIaH on cis-elements similar to those found in contractile protein genes suggests that the striated muscle-specific expression is coregulated by mechanisms that control the lineage-specific expression of several contractile and cytosolic proteins.

Non-NASA Center↗

Some growth factors stimulate cultured adult rabbit ventricular myocyte hypertrophy in the absence of mechanical loading

Cultured adult rabbit cardiac myocytes treated with recombinant growth factors display enhanced rates of protein accumulation (ie, growth) in response to insulin and insulin-like growth factors (IGFs), but epidermal growth factor, acidic or basic fibroblast growth factor, and platelet-derived growth factor failed to increase contractile protein synthesis or growth of the heart cells. Insulin and IGF-1 increased growth rates by stimulating anabolic while simultaneously inhibiting catabolic pathways, whereas IGF-2 elevated growth modestly by apparently inhibiting lysosomal proteolysis. Neutralizing antibodies directed against either IGF-1 or IGF-2 or IGF binding protein 3 blocked protein accumulation. A monoclonal antibody directed against the IGF-1 receptor also inhibited changes in protein turnover provoked by recombinant human IGF-1 but not IGF-2. Of the other growth factors tested, only transforming growth factor-beta 1 increased the fractional rate of myosin heavy chain (MHC) synthesis, with beta-MHC synthesis being elevated and alpha-MHC synthesis being suppressed. However, the other growth factors were able to modestly stimulate the rate of DNA synthesis in this preparation. Bromodeoxyuridine labeling revealed that these growth factors increased DNA synthesis in myocytes and nonmyocytes alike, but the heart cells displayed neither karyokinesis or cytokinesis. In contrast, cocultures of cardiac myocytes and nonmyocytes and nonmyocyte-conditioned culture medium failed to enhance the rate of cardiac MHC synthesis or its accumulation, implying that quiescent heart cells do not respond to "conditioning" by cardiac nonmyocytes. These findings demonstrated that insulin and the IGFs promote passively loaded cultured adult rabbit heart cells to hypertrophy but suggest that other growth factors tested may be limited in this regard.

NASA Discipline Cell Biology↗

Glucocorticoid-induced skeletal muscle atrophy in vitro is attenuated by mechanical stimulation

Glucocorticoids induce rapid atrophy of fast skeletal myofibers in vivo, and either weight lifting or endurance exercise reduces this atrophy by unknown mechanisms. We examined the effects of the synthetic glucocorticoid dexamethasone (Dex) on protein turnover in tissue-cultured avian fast skeletal myofibers and determined whether repetitive mechanical stretch altered the myofiber response to Dex. In static cultures after 3-5 days, 10(-8) M Dex decreased total protein content 42-74%, total protein synthesis rates 38-56%, mean myofiber diameter 35%, myosin heavy chain (MHC) content 86%, MHC synthesis rate 44%, and fibronectin synthesis rate 29%. Repetitive 10% stretch-relaxations of the cultured myofibers for 60 s every 5 min for 3-4 days prevented 52% of the Dex-induced decrease in protein content, 42% of the decrease in total protein synthesis rate, 77% of the decrease in MHC content, 42% of the decrease in MHC synthesis rate, and 67% of the decrease in fibronectin synthesis rate. This in vitro model system will complement in vivo studies in understanding the mechanism by which mechanical activity and glucocorticoids interact to regulate skeletal muscle growth.

Non-NASA Center↗

Fluorescent Approaches to High Throughput Crystallography

We have shown that by covalently modifying a subpopulation, less than or equal to 1%, of a macromolecule with a fluorescent probe, the labeled material will add to a growing crystal as a microheterogeneous growth unit. Labeling procedures can be readily incorporated into the final stages of purification, and the presence of the probe at low concentrations does not affect the X-ray data quality or the crystallization behavior. The presence of the trace fluorescent label gives a number of advantages when used with high throughput crystallizations. The covalently attached probe will concentrate in the crystal relative to the solution, and under fluorescent illumination crystals show up as bright objects against a dark background. Non-protein structures, such as salt crystals, will not incorporate the probe and will not show up under fluorescent illumination. Brightly fluorescent crystals are readily found against less bright precipitated phases, which under white light illumination may obscure the crystals. Automated image analysis to find crystals should be greatly facilitated, without having to first define crystallization drop boundaries as the protein or protein structures is all that shows up. Fluorescence intensity is a faster search parameter, whether visually or by automated methods, than looking for crystalline features. We are now testing the use of high fluorescence intensity regions, in the absence of clear crystalline features or "hits", as a means for determining potential lead conditions. A working hypothesis is that kinetics leading to non-structured phases may overwhelm and trap more slowly formed ordered assemblies, which subsequently show up as regions of brighter fluorescence intensity. Preliminary experiments with test proteins have resulted in the extraction of a number of crystallization conditions from screening outcomes based solely on the presence of bright fluorescent regions. Subsequent experiments will test this approach using a wider range of proteins. The trace fluorescently labeled crystals will also emit with sufficient intensity to aid in the automation of crystal alignment using relatively low cost optics, further increasing throughput at synchrotrons.

Pusey, Marc L.↗

Dynamically controlled crystal growth system

Crystal growth can be initiated and controlled by dynamically controlled vapor diffusion or temperature change. In one aspect, the present invention uses a precisely controlled vapor diffusion approach to monitor and control protein crystal growth. The system utilizes a humidity sensor and various interfaces under computer control to effect virtually any evaporation rate from a number of different growth solutions simultaneously by means of an evaporative gas flow. A static laser light scattering sensor can be used to detect aggregation events and trigger a change in the evaporation rate for a growth solution. A control/follower configuration can be used to actively monitor one chamber and accurately control replicate chambers relative to the control chamber. In a second aspect, the invention exploits the varying solubility of proteins versus temperature to control the growth of protein crystals. This system contains miniature thermoelectric devices under microcomputer control that change temperature as needed to grow crystals of a given protein. Complex temperature ramps are possible using this approach. A static laser light scattering probe also can be used in this system as a non-invasive probe for detection of aggregation events. The automated dynamic control system provides systematic and predictable responses with regard to crystal size. These systems can be used for microgravity crystallization projects, for example in a space shuttle, and for crystallization work under terrestial conditions. The present invention is particularly useful for macromolecular crystallization, e.g. for proteins, polypeptides, nucleic acids, viruses and virus particles.

Bray, Terry L.↗

Nutrition for Space Exploration

Nutrition has proven to be critical throughout the history of human exploration, on both land and water. The importance of nutrition during long-duration space exploration is no different. Maintaining optimal nutritional status is critical for all bodily systems, especially in light of the fact that that many are also affected by space flight itself. Major systems of concern are bone, muscle, the cardiovascular system, the immune system, protection against radiation damage, and others. The task ahead includes defining the nutritional requirements for space travelers, ensuring adequacy of the food system, and assessing crew nutritional status before, during, and after flight. Accomplishing these tasks will provide significant contributions to ensuring crew health on long-duration missions. In addition, development and testing of nutritional countermeasures to effects of space flight is required, and assessment of the impact of other countermeasures (such as exercise and pharmaceuticals) on nutrition is also critical for maintaining overall crew health. Vitamin D stores of crew members are routinely low after long-duration space flight. This occurs even when crew members take vitamin D supplements, suggesting that vitamin D metabolism may be altered during space flight. Vitamin D is essential for efficient absorption of calcium, and has numerous other benefits for other tissues with vitamin D receptors. Protein is a macronutrient that requires additional study to define the optimal intake for space travelers. Administration of protein to bed rest subjects can effectively mitigate muscle loss associated with disuse, but too much or too little protein can also have negative effects on bone. In another bed rest study, we found that the ratio of protein to potassium was correlated with the level of bone resorption: the higher the ratio, the more bone resorption. These relationships warrant further study to optimize the beneficial effect of protein on both bone and muscle during space flight. Omega3 fatty acids are currently being studied as a means of protecting against radiation-induced cancer. They have also recently been implicated as having a role in mitigating the physical wasting, or cachexia, caused by cancer. The mechanism of muscle loss associated with this type of cachexia is similar to the mechanism of muscle loss during disuse or space flight. Omega3 fatty acids have already been shown to have protective effects on bone and cardiovascular function. Omega3 fatty acids could be an ideal countermeasure for space flight because they have protective effects on multiple systems. A definition of optimal nutrient intake requirements for long-duration space travel should also include antioxidants. Astronauts are exposed to numerous sources of oxidative stress, including radiation, elevated oxygen exposure during extravehicular activity, and physical and psychological stress. Elevated levels of oxidative damage are related to increased risk for cataracts, cardiovascular disease, and cancer. Many groundbased studies show the protective effects of antioxidants against oxidative damage induced by radiation or oxygen. Balancing the diet with foods that have high levels of antioxidants would be another ideal countermeasure because it should have minimal side effects on crew health. Antioxidant supplements, however, are often used without having data on their effectiveness or side effects. High doses of supplements have been associated with bone and cardiovascular problems, but research on antioxidant effects during space flight has not been conducted. Much work must be done before we can send crews on exploration missions. Nutrition is often assumed to be the simple provision of food items that will be stable throughout the mission. As outlined briefly above, the situation is much more complex than food provision. As explorers throughout history have found, failure to truly understand the role of nutrition can be catastrophic. When huns are in environments unlike any they have seen before, this is more true than ever.

Smith, Scott M.↗

Nutrition Research: Basis for Station Requirements

Prior to the Shuttle program, all understanding of nutritional needs in space came from Skylab metabolic research. Because Shuttle flights were short, most less than 14 days, research focused on major nutritional issues: energy (calories), protein and amino acids, water and electrotypes, with some more general physiology studies that related to iron and calcium. Using stable isotope tracer studies and diet intake records, we found that astronauts typically did not consume adequate calories to meet energy expenditure. To monitor energy and nutrient intake status and provide feedback to the flight surgeon and the astronauts, the International Space Station (ISS) program implemented a weekly food frequency questionnaire and routine body mass measurements. Other Shuttle investigations found that protein turnover was higher during flight, suggesting there was increased protein degradation and probably concurrent increase in protein synthesis, and this occurred even in cases of adequate protein and caloric intake. These results may partially explain some of the loss of leg muscle mass. Fluid and electrolyte flight studies demonstrated that water intake, like energy intake, was lower than required. However, sodium intakes were elevated during flight and likely related to other concerns such as calcium turnover and other health-related issues. NASA is making efforts to have tasty foods with much lower salt levels to reduce sodium intake and to promote fluid intake on orbit. Red blood cell studies conducted on the Shuttle found decreased erythrogenesis and increased serum ferritin levels. Given that the diet is high in iron there may be iron storage health concerns, especially related to the role of iron in oxidative damage, complicated by the stress and radiation. The Shuttle nutrition research lead to new monitoring and research on ISS. These data will be valuable for future NASA and commercial crewed missions.

Lane, Helen W.↗

Use of Aquaporins to Achieve Needed Water Purity On ISS for the EMU Space Suit System

With the U.S. Space Shuttle fleet retired, the supply of extremely high-quality water 'super-Q' - required for the EMU Space suit cooling on this ISS - will become a significant operational hardware challenge in the very near future. A proposed potential solution is the use of a filtration system consisting of a semi-permeable membrane embedded with aquaporin proteins. Aquaporins are a special class of trans-membrane proteins that facilitate passive transport of water and other substances across a membrane. The specificity of these proteins is such that only water is allowed through the protein structure, and this novel property invites their adaptation for use in water filtration systems, specifically usage on the ISS for the EMU space suit system. These proteins are found in many living systems and have been developed for commercial use today.

Hill, Terry R.↗

Relationships Between Seed Weight, Germination Potential and Biochemical Reserves of Maritime Pine in Morocco: Elements for Tree Seedlings Improvement

Selection of quality seeds in breeding programs can significantly improve seedling productivity. Germination and biochemical analyses on seeds from ten natural populations of maritime pine (Pinus pinaster Ait.) in Morocco reveals significant differences among populations in seed weight, germination characters and protein content in both dry seeds and megagametophytes. During germination, the mobilization of protein content in megagametophyte is significantly different among populations than sugar content. A strong positive correlation between the germination capacity and the protein content in both dry seeds and megagametophytes indicates that the best populations in term of germination capacity may also be the richest in protein content. The present study finds that seed weight is not a good indicator for quality seed selection, nor is it recommended to increase the degree of germinability. Our results suggest that the pine population in southern Morocco might have adapted to drought conditions as it is characterized by heavy seed weight and lower speed of protein content mobilization in megagametophyte compared to northern populations growing in temperate climate.

Wahid, Nadya↗

Microwave Tissue Soldering for Immediate Wound Closure

A novel approach for the immediate sealing of traumatic wounds is under development. A portable microwave generator and handheld antenna are used to seal wounds, binding the edges of the wound together using a biodegradable protein sealant or solder. This method could be used for repairing wounds in emergency settings by restoring the wound surface to its original strength within minutes. This technique could also be utilized for surgical purposes involving solid visceral organs (i.e., liver, spleen, and kidney) that currently do not respond well to ordinary surgical procedures. A miniaturized microwave generator and a handheld antenna are used to deliver microwave energy to the protein solder, which is applied to the wound. The antenna can be of several alternative designs optimized for placement either in contact with or in proximity to the protein solder covering the wound. In either case, optimization of the design includes the matching of impedances to maximize the energy delivered to the protein solder and wound at a chosen frequency. For certain applications, an antenna could be designed that would emit power only when it is in direct contact with the wound. The optimum frequency or frequencies for a specific application would depend on the required depth of penetration of the microwave energy. In fact, a computational simulation for each specific application could be performed, which would then match the characteristics of the antenna with the protein solder and tissue to best effect wound closure. An additional area of interest with potential benefit that remains to be validated is whether microwave energy can effectively kill bacteria in and around the wound. Thus, this may be an efficient method for simultaneously sterilizing and closing wounds.

Arndt, G. Dickey↗

Processes that Drove the Transition from Chemistry to Biology: Concepts and Evidence

Two properties are particularly germane to the transition from chemistry to biology. One is the emergence of complex molecules (polymers) capable of performing non-trivial functions, such as catalysis, energy transduction or transport across cell walls. The other is the ability of several functions to work in concert to provide reproductive advantage to systems hosting these functions. Biological systems exhibit these properties at remarkable levels of efficiency and accuracy in a way that appears effortless. However, dissection of these properties reveals great complexities that are involved. This opens a question: how a simple, ancestral system could have acquired the required properties? Other questions follow. What are the chances that a functional polymer emerges at random? What is the minimum structural complexity of a polymer to carry out a function at a reasonable level of efficiency? Can we identify concrete, protobiologically plausible mechanisms that yield advantageous coupling between different functions? These and similar questions are at the core of the main topic of this session: how soulless chemistry became life? Clearly, we do not have complete answers to any of these questions. However, in recent years a number of new and sometimes unexpected clues have been brought to light. Of particular interest are proteins because they are the main functional polymers in contemporary cells. The emergence of protein functions is a puzzle. It is widely accepted that a well ]defined, compact structure (fold) is a prerequisite for function. It is equally widely accepted that compact folds are rare among random amino acid polymers. Then, how did protein functionality start? According to one hypothesis well folded were preceded by their poorly folded, yet still functional ancestors. Only recently, however, experimental evidence supporting this hypothesis has been presented. In particular, a small enzyme capable of ligating two RNA fragments with the rate of 106 above background was evolved in vitro. This enzyme does not look like any contemporary protein. It is very flexible and its structure is kept together just by a single salt bridge between a charged residue and a coordinating zinc. A similar picture emerges from studies of simple transmembrane channels that mimic those in ancestral cells. Again, they are extremely flexible and do not form a conventional pore. Yet, they efficiently mediate ion transport. Studies on simple proteins that are on-going in several laboratories hold promise of revealing crucial links between chemical and biological catalysis and other ubiquitous cell functions. Interaction between composition, growth and division of protobiologically relevant vesicles and metabolic reactions that they encapsulate is an example of coupling between simple functions that promotes reproduction and evolution. Recent studies have demonstrated possible mechanisms by which vesicles might have evolved their composition from fatty acids to phospholipids, thus facilitating a number of new cellular functions. Conversely, it has been also demonstrated that an encapsulated metabolism might drive vesicle division. These are, again, examples of processes that might have driven the transition from chemistry to biology.

Pohorille, Andrew↗

Nanoscale Surface Plasmonics Sensor With Nanofluidic Control

Conventional quantitative protein assays of bodily fluids typically involve multiple steps to obtain desired measurements. Such methods are not well suited for fast and accurate assay measurements in austere environments such as spaceflight and in the aftermath of disasters. Consequently, there is a need for a protein assay technology capable of routinely monitoring proteins in austere environments. For example, there is an immediate need for a urine protein assay to assess astronaut renal health during spaceflight. The disclosed nanoscale surface plasmonics sensor provides a core detection method that can be integrated to a lab-on-chip device that satisfies the unmet need for such a protein assay technology. Assays based upon combinations of nanoholes, nanorings, and nanoslits with transmission surface plasmon resonance (SPR) are used for assays requiring extreme sensitivity, and are capable of detecting specific analytes at concentrations as low as picomole to femtomole level in well-controlled environments. The device operates in a transmission mode configuration in which light is directed at one planar surface of the array, which functions as an optical aperture. The incident light induces surface plasmon light transmission from the opposite surface of the array. The presence of a target analyte is detected by changes in the spectrum of light transmitted by the array when a target analyte induces a change in the refractive index of the fluid within the nanochannels. This occurs, for example, when a target analyte binds to a receptor fixed to the walls of the nanochannels in the array. Independent fluid handling capability for individual nanoarrays on a nanofluidic chip containing a plurality of nanochannel arrays allows each array to be used to sense a different target analyte and/or for paired arrays to analyze control and test samples simultaneously in parallel. The present invention incorporates transmission mode nanoplasmonics and nanofluidics into a single, microfluidically controlled device. The device comprises one or more arrays of aligned nanochannels that are in fluid communication with inflowing and outflowing fluid handling manifolds that control the flow of fluid through the arrays. The array acts as an aperture in a plasmonic sensor. Fluid, in the form of a liquid or a gas and comprising a sample for analysis, is moved from an inlet manifold through the nanochannel array, and out through an exit manifold. The fluid may also contain a reagent used to modify the interior surfaces of the nanochannels, and/or a reagent required for the detection of an analyte.

Wei, Jianjun↗

Use of Aquaporins to Achieve Needed Water Purity On ISS for the EMU Space Suit System

Use of Aquaporins to Achieve Needed Water Purity On ISS for the EMU Space Suit System. With the U.S. Space Shuttle fleet retired, the supply of extremely high-quality water "super-Q" - required for the EMU Space suit cooling on this ISS - will become a significant operational hardware challenge in the very near future. A proposed potential solution is the use of a filtration system consisting of a semi-permeable membrane embedded with aquaporin proteins. Aquaporins are a special class of trans-membrane proteins that facilitate passive transport of water and other substances across a membrane. The specificity of these proteins is such that only water is allowed through the protein structure, and this novel property invites their adaptation for use in water filtration systems, specifically usage on the ISS for the EMU space suit system. These proteins are found in many living systems and have been developed for commercial use today.

Hill, Terry↗

Carbon Nanostructures in Biological Sensors for Space and Terrestrial Applications

Biosensing devices comprised of carbon nanotubes and nanofibers have been developed for astronaut crew point-of-care. Their inherent nanometer scale, high conductivity, wide potential window, good biocompatibility and well-defined surface chemistry make them ideal candidates as biosensor electrodes. Here, we report two studies using carbon nanotube and carbon nanofiber electrodes for biomedical applications. First, a 3x3 electrode device, with each electrode containing 40,000 carbon nanofiber nanoelectrodes was fabricated on silicon using traditional microfabrication processing. The device was demonstrated as a multiplexed immunosensor for simultaneous, label-free detection of cardiac troponin-I, C-reactive protein and myoglobin. Antibodies specific to cardiac troponin-I, C-reactive protein and myoglobin were covalently bound to the CNF surface and were characterized using electrochemical impedance spectroscopy and differential pulse voltammetry. Each step of the modification process resulted in changes in resistance to charge transfer due to the changes at the electrode surface upon antibody immobilization and binding to the specific cardiac protein. The real-time label free detection of the three cardiac markers from pure components and mixtures was demonstrated with high sensitivity, down to 0.2 ng/mL, and good selectivity. Detection in human blood serum did not present false positives from non-specific protein adsorption. More recently, this detection scheme has been applied to inkjet printed carbon nanotube electrodes on Kapton and paper. Printed devices have several unique advantages including simple and inexpensive fabrication. The results demonstrate that these sensors can serve a miniaturized, low cost device for detection of proteins in complex mixtures making this platform a good candidate for early stage diagnosis of myocardial infarction. Future inkjet printed devices can be fabricated have the added advantage in their suitability to be manufactured in an in-space, microgravity environment.

carbon nanotubes↗