Die vorliegende Arbeit wurde von der Europäischen Union IST-Programm (IST-2002-507464-Projekt &quot;- - A Virtual Reality Intelligent Multi-Sensor Wearable-System für Phobien&quot; Treatment INTREPID &quot;) unterstützt.

Das klinische Protokoll für die Behandlung der generalisierten Angststörung (GAD) ist gleichzeitig innerhalb und außerhalb der Praxis des Therapeuten programmiert. Relax-Programm ist eine Kombination aus &quot;Progressive Muskelentspannung Training&quot; (PMRT), dem klassischen Verfahren von Jacobsen beschrieben, dass Anspannung und Entspannung verschiedener Muskelgruppen bei Kenntnisnahme der Kontrast Empfindungen und &quot;Guided Imagery&quot;-Programm beinhaltet. In unserem Training Bildern wird durch die visuellen Hinweis von einem erholsamen virtuellen Umgebung ersetzt. INSIDE der Praxis des Therapeuten. Patienten teilnehmen zu 8 Trainingseinheiten mit dem Therapeuten, in 4 Wochen (2 Sitzungen pro Woche) verteilt. Jede Sitzung beginnt und endet mit einer klinischen Beurteilung, dass die Maßnahme der die Angst des Patienten gibt. Plan der Sitzungen in der Praxis des Therapeuten: <ol><li> Erste klinische Evaluierung der Zustand des Patienten; </li><li> Der Patient wird mit Biosensoren, dass seine / ihre physiologische Parameter (Hautleitwert, Herzfrequenz, Atmung) Aufzeichnung verbunden. Eine Baseline-Messung dieser Parameter wird für 3 Minuten registriert; </li><li> Der Patient trägt ein Head-Mounted-Display mit einem PC verbunden und wickelt einen Joystick; </li><li> Der Patient beginnt die Erkundung der virtuellen Umgebung: einer wunderschönen tropischen Insel mit Blick auf den Ozean. Der Patient, nach der Erzählung von dem Therapeuten aufgenommen, erreicht die Insel mit dem Boot und erforschen. Nach einem Fußweg, führt ihm / ihr durch die Insel, kommt der Patient zum Ausgangspunkt, wo verschiedene Tafeln geben die Richtungen der verschiedenen Zielgebiete. In jedem dieser Bereiche eine entspannende Übung vorgesehen ist; während dieser Ausbildung, folgen Sie der Beschilderung durch die Stimme-guide angegeben wird, versucht der Patient von ihm / ihr zu entspannen, während einige Elemente der virtuellen Umgebung direkt durch seine modifizierte / ihre physiologische Parameter erfasst in Echtzeit. Somit erhält der Patient eine sofortige Rückmeldung seiner / ihrer Ebene der Aktivierung (wie in der traditionellen Biofeedback-Techniken), aber mit dem zusätzlichen Wert durch die virtuelle Umgebung, die er / sie ist immersiv erkunden gegeben. Die Zielgebiete sind gebunden: <ul><li> Campfire: physiologische Parameter steuern das Feuer Intensität, so dass die Reduzierung des Patienten physiologische Aktivierung Ergebnisse auf eine Reduzierung der Feuer, bis es erlischt; </li><li> Waves: physiologische Parameter steuern die Wellen, bis das Meer sich ganz ruhig; </li><li> Wasserfall: die Reduktion der physiologischen Erregung entspricht einer Reduktion des Strom-Intensität, bis das Wasser aufhört zu fallen. </li><li> Gazebo: anders als die anderen, ist diese Übung kundengerecht durch den Patienten, die Worte oder Bilder im Zusammenhang mit ihm / ihr persönliche belastenden Ereignissen wählen können. Diese Technik dient als Stressimpfung Programm, und der Patient kann die Verkleinerung-Fading-floating des ausgewählten Artikels Erfahrung - je nach seiner / ihrer Fähigkeit, ihn / sie sich entspannen; </li></ul></li><li> Nach der Fertigstellung der Virtual-Reality-Sitzung werden physiologische Parameter wieder in Ruhe aufgezeichnet; </li><li> Abschließende klinische Evaluation der Zustand des Patienten. </li></ol> OUTSIDE Praxis des Therapeuten: Um die Wirksamkeit der Ausbildung zu verbessern und die Auswirkungen der Entspannung zu erhöhen, die Patienten Entspannungstechniken außerhalb Praxis des Therapeuten Praxis erleben eine Homecare-Szenario. Dies ist durch Vorlage der gleichen virtuellen Umgebung während der Therapie auf einem mobilen Gerät erfahrenen realisiert. Das mobile Gerät wird via Bluetooth auf einen Satz von Biosensoren, die Herzfrequenz und die Hautleitfähigkeit umfasst verbunden. Dieses Gerät ist klein genug und leicht zu benutzen, um selbstständig durch den Patienten behandelt werden (siehe Abb. 1).. Der Patient ist erforderlich, um Entspannung Fähigkeiten mindestens einmal am Tag trainieren für die gesamte Dauer der Behandlung (ein Monat), im Anschluss an die Behandlung zu planen, indem der Therapeut zur Verfügung gestellt. Repräsentative Ergebnisse Da beide Biofeedback und virtuelle Realität bekannt sind, um wirksam zur Entspannung zu induzieren, ist die wichtigste erwartete Ergebnis, dass eine Kombination dieser beiden Methoden effektiver reduzieren Angst bei Patienten mit generalisierter Angststörung betroffen, als die beiden Techniken einzeln verwendet wird. <img src="/files/ftp_upload/1554/Fig1.jpg" alt="Abbildung 1" /> Abbildung 1: GSR / HR Biosensoren Modul.

<ol>
	<li>Arntz, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cognitive+therapy+versus+applied+relaxation+as+treatment+of+generalized+anxiety+disorder.">Cognitive therapy versus applied relaxation as treatment of generalized anxiety disorder.</a> Behav Res Ther. 41 (6), 633-633 (2003).</li><li>Siev, J., Chambless, D. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Specificity+of+treatment+effects:+cognitive+therapy+and+relaxation+for+generalized+anxiety+and+panic+disorders.">Specificity of treatment effects: cognitive therapy and relaxation for generalized anxiety and panic disorders.</a> Journal of consulting and clinical psychology. 75 (4), 513-513 (2007).</li><li>Jacobsen, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Progressive relaxation. , (1938).</li><li>Villani, D., Riva, F., Riva, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=New+technologies+for+relaxation:+the+role+of+presence.">New technologies for relaxation: the role of presence.</a> International journal of Stress Management. 14 (3), 260-260 (2007).</li></ol>

The authors have nothing to disclose.

Unsere Hypothese ist, dass die Kombination aus virtueller Realität und Biofeedback, das direkt modifiziert die virtuelle Umgebung, verglichen mit der virtuellen Realität werden allein in eine bessere und schnellere Erholung Lernen führen. Wenn dem so ist, hat dieses neue Instrument, um Angstzustände könnten angewendet und getestet werden, auch in anderen Angst zusammenhängenden Pathologien.

the use of biofeedback in clinical virtual reality: the intrepid project

Generalisierte Angststörung (GAD) ist eine psychiatrische Erkrankung, die durch eine konstante und unspezifische Angst, die mit Alltagsaktivitäten behindert aus. Die hohe Prävalenz in der allgemeinen Bevölkerung und die starken Einschränkungen sie verursacht, auf die Notwendigkeit, neue effiziente Strategien zu ihrer Behandlung zu finden. Zusammen mit den kognitiv-verhaltenstherapeutischen Behandlungen, stellt Entspannung ein nützlicher Ansatz für die Behandlung von GAD, aber es hat die Einschränkung, dass es schwer zu erlernen. Die INTREPID Projekt zielt darauf ab, ein neues Instrument, um Angst-Erkrankungen zu behandeln und seine klinische Wirksamkeit bei der Verringerung der Angst-Symptome Tests zu implementieren. Die Innovation dieses Ansatzes ist die Kombination aus virtueller Realität und Biofeedback, so dass der erste direkt vom Ausgang des zweiten geändert. Auf diese Weise wird der Patient aufmerksam gemacht seine Reaktionen durch die Modifikation einiger Features der VR-Umgebung in Echtzeit. Mit mentalen Übungen lernt der Patient auf diese physiologischen Parameter steuern und mit dem Feedback von der virtuellen Umgebung zur Verfügung gestellt in der Lage ist, seine oder ihre Erfolge zu messen. Der ergänzende Einsatz von tragbaren Geräten wie PDA oder Smart-Phones, kann der Patient zu Hause durchführen, individuell und eigenständig, die gleichen Übungen in Praxis des Therapeuten erlebt. Das Ziel ist es, die gelernt Protokoll in einer realen Leben Kontext zu verankern, so die Verbesserung der Patienten die Fähigkeit, mit ihren Symptomen zu befassen. Das erwartete Ergebnis ist eine bessere und schnellere Erlernen von Entspannungstechniken, und damit eine erhöhte Wirksamkeit der Behandlung, wenn sie mit herkömmlichen klinischen Protokolle verglichen.

Watch this Scientific Journal Video about The use of Biofeedback in Clinical Virtual Reality: The INTREPID Project at JoVE.com

The use of Biofeedback in Clinical Virtual Reality: The INTREPID Project

Das Project on Virtual Reality Intelligent Multi-Sensor-System für die Behandlung von Angst-Erkrankungen (Intrepid) ist auf die Entwicklung eines Multi-Sensor-kontextsensitive Virtual-Reality-System für die Behandlung von Angst-Erkrankungen ausgerichtet.

Der Einsatz von Biofeedback in der klinischen Virtual Reality: Die INTREPID Projekt

Generalized anxiety disorder (GAD) is a psychiatric disorder characterized by a constant and unspecific anxiety that interferes with daily-life ...

the-use-biofeedback-clinical-virtual-reality-intrepid

Research

JoVE Journal

Biology

15.0K Aufrufe.  Istituto Auxologico Italiano. Das Project on Virtual Reality Intelligent Multi-Sensor-System für die Behandlung von Angst-Erkrankungen (Intrepid) ist auf die Entwicklung eines Multi-Sensor-kontextsensitive Virtual-Reality-System für die Behandlung von Angst-Erkrankungen ausgerichtet.

Serie: The use of Biofeedback in Clinical Virtual Reality: The INTREPID Project

Use of Rotorod as a Method for the Qualitative Analysis of Walking in Rat

High speed videoanalysis of the details of movement can provide a source of information about qualitative aspects of walking movements. When walking on a rotorod, animals remain in approximately the same place making repetitive movements of stepping. Thus the task provides a rich source of information on the details of foot stepping movements. Subjects were hemi-Parkinson analogue rats, produced by injection of 6-hydroxydopamine (6-OHDA) into the right nigrostriatal bundle to deplete nigrostriatal dopamine (DA). The present report provides a video analysis illustration of animals previously were filmed from frontal, lateral, and posterior views as they walked (15). Rating scales and frame-by-frame replay of the video records of stepping behavior indicated that the hemi-Parkinson rats were chronically impaired in posture and limb use contralateral to the DA-depletion. The contralateral limbs participated less in initiating and sustaining propulsion than the ipsilateral limbs. These deficits secondary to unilateral DA-depletion show that the rotorod provides a use task for the analysis of stepping movements.

The rotorod test is used to assess motor status in the walking movement of hemi-Parkinson analogue rats.

Verwendung von Drehstabtest als Methode für die qualitative Analyse von Walking in Rat

High speed videoanalysis of the details of movement can provide a source of information about qualitative aspects of walking movements. When walking on a ...

Forschung

Biologie

Psychophysiological Stress Assessment Using Biofeedback 

In the last half century, research in biofeedback has shown the extent to which the human mind can influence the functioning of the autonomic nervous system, previously thought to be outside of conscious control. By letting people observe signals from their own bodies, biofeedback enables them to develop greater awareness of their physiological and psychological reactions, such as stress, and to learn to modify these reactions. Biofeedback practitioners can facilitate this process by assessing people s reactions to mildly stressful events and formulating a biofeedback-based treatment plan. During stress assessment the practitioner first records a baseline for physiological readings, and then presents the client with several mild stressors, such as a cognitive, physical and emotional stressor. Variety of stressors is presented in order to determine a person's stimulus-response specificity, or differences in each person's reaction to qualitatively different stimuli. This video will demonstrate the process of psychophysiological stress assessment using biofeedback and present general guidelines for treatment planning.

Psychophysiological Stress Assessment Using Biofeedback

In this video we will describe one method of assessing person's psychophysiological reaction to stress using biofeedback. We will also present general guidelines for treatment planning.

Psychophysiologische Stress Assessment mit Biofeedback

In the last half century, research in biofeedback has shown the extent to which the human mind can influence the functioning of the autonomic nervous ...

Use of the Protease Fluorescent Detection Kit to Determine Protease Activity

The Protease Fluorescent Detection Kit provides ready-to-use reagents for detecting the presence of protease activity. This simple assay to detect protease activity uses casein labeled with fluorescein isothiocyanate (FITC) as the substrate.
Protease activity results in the cleavage of the FITC-labeled casein substrate into smaller fragments, which do not precipitate under acidic conditions. After incubation of the protease sample and substrate, the reaction is acidified with the addition of trichloroacetic acid (TCA). The mixture is then centrifuged with the undigested substrate forming a pellet and the smaller, acid soluble fragments remaining in solution. The supernatant is neutralized and the fluorescence of the FITC-labeled fragments is measured.
The described kit procedure detects the trypsin protease control at a concentration of approximately 0.5 &mu;g/ml (5 ng of trypsin added to the assay). This sensitivity can be increased with a longer incubation time, up to 24 hours. The assay is performed in microcentrifuge tubes and procedures are provided for fluorescence detection using either cuvettes or multiwell plates.

The Protease Fluorescent Detection Kit is designed for the measurement of protease activity using fluorometry. It is also suitable for detection of trace amounts of protease contamination. The method is based on the proteolytic hydroysis of a proprietary formulation of a FITC-labeled casein substrate.

Die Nutzung der Protease Fluorescent Detection Kit für Protease-Aktivität zu bestimmen

The Protease Fluorescent Detection Kit provides ready-to-use reagents for detecting the presence of protease activity. This simple assay to detect ...

The use of SC1 (Pluripotin) to Support mESC Self-renewal in the Absence of LIF

Mouse embryonic stem (ES) cells are conventionally cultured with Leukemia Inhibitory Factor (LIF) to maintain self-renewal.1 However, LIF is expensive and activation of the LIF/JAK/STAT3 pathway is not absolutely required to maintain the self-renewal state.2 The SC1 small molecule may be an economical alternative to LIF. SC1 functions through dual inhibition of Ras-GAP and ERK1.3 Illustration of its mechanism of action makes it a useful tool to study the fundamental molecular mechanism of self-renewal. Here we demonstrate the procedure for culturing mouse ES cells in the presence of SC1 and show that they are able to maintain self-renewal in the absence of LIF. Cells cultured with SC1 showed similar morphology compared to cells maintained with LIF. Both exhibited typical mouse ES morphology after five passages. Expression of typical pluripotency markers (Oct4, Sox2, Nanog, and SSEA1) was observed after five passages in the presence of SC1. Furthermore, SC1 caused no overt toxicity on mouse ES cells.

The use of SC1 Pluripotin to Support mESC Self-renewal in the Absence of LIF

SC1 functions through dual inhibition of Ras- GAP and ERK1. We tested the function of SC1 in supporting mouse ES cell self-renewal in the absence of LIF and showed that SC1 is able to maintain self-renewal of mouse ES cell cultures.

Die Verwendung von SC1 (Pluripotin) zu MESC Self-Erneuerung in der Abwesenheit von LIF-Support

Mouse embryonic stem (ES) cells are conventionally cultured with Leukemia Inhibitory Factor (LIF) to maintain self-renewal.1 However, LIF is expensive and ...

Purification of the M. magneticum Strain AMB-1 Magnetosome Associated Protein MamA&Delta;41

Magnetotactic bacteria comprise a diverse group of aquatic microorganisms that are able to orientate themselves along geomagnetic fields. This behavior is believed to aid their search for suitable environments (1). This capability is conferred by the magnetosome, a subcellular organelle that consists of a linear-chain assembly of lipid vesicles each able to biomineralize and enclose a ~50-nm crystal of magnetite or greigite. A principle component of the magnetosome that was shown to be required for the formation of functional vesicles is MamA. MamA is a highly abundant magnetosome-associated protein which is one of the most characterized magnetosome-associated proteins in vivo (2-6). This article focuses on the purification of MamA, which despite being studied in vivo, no clear functional or structural details have been identified for it. Bioinformatics analysis suggested that MamA is a tetra-tricopeptide repeat (TPR) containing protein. TPR is a structural motif found as such or forming part of a bigger fold in a wide range of proteins, it serves as a template for protein-protein interactions and mediates multi-protein complexes (7). TPRs are involved in many crucial tasks in eukaryotic cell organelle processes and many bacterial pathways (8-14). In order to understand MamA, a unique TPR containing protein, highly purified protein is required as a first step. In this article, we present the purification protocol for a stable MamA deletion mutant (MamA&Delta;41) from M. magneticum AMB-1.

Purification of the &lt;em&gt;M. magneticum&lt;/em&gt; Strain AMB-1 Magnetosome Associated Protein MamA&amp;Delta;41

MamA is a unique Magnetosome associated protein which was shown to be involved in magnetosome activation. Here we present the purification protocol of MamA deletion mutant (MamA&Delta;41) from M. magneticum AMB-1.

Die Reinigung des M. magneticum Stamm AMB-1 Magnetosom assoziierten Protein MamAΔ41

Magnetotactic bacteria comprise a diverse group of aquatic microorganisms that are able to orientate themselves along geomagnetic fields. This behavior is ...

Live Imaging of Apoptotic Cell Clearance during Drosophila Embryogenesis

The proper elimination of unwanted or aberrant cells through apoptosis and subsequent phagocytosis (apoptotic cell clearance) is crucial for normal development in all metazoan organisms. Apoptotic cell clearance is a highly dynamic process intimately associated with cell death; unengulfed apoptotic cells are barely seen in vivo under normal conditions. In order to understand the different steps of apoptotic cell clearance and to compare 'professional' phagocytes - macrophages and dendritic cells to 'non-professional' - tissue-resident neighboring cells, in vivo live imaging of the process is extremely valuable. Here we describe a protocol for studying apoptotic cell clearance in live Drosophila embryos. To follow the dynamics of different steps in phagocytosis we use specific markers for apoptotic cells and phagocytes. In addition, we can monitor two phagocyte systems in parallel: 'professional' macrophages and 'semi-professional' glia in the developing central nervous system (CNS). The method described here employs the Drosophila embryo as an excellent model for real time studies of apoptotic cell clearance.

Live Imaging of Apoptotic Cell Clearance during Drosophila Embryogenesis

Here we describe an effective method for studying dynamics of apoptotic cell clearance in vivo. This method employs live Drosophila embryos as a powerful model for monitoring phagocytosis of apoptotic cells using specific labeling of apoptotic cells and phagocytes.

Live-Imaging von apoptotischen Ausverkauf während<em&gt; Drosophila</em&gt; Embryogenese

The  proper elimination of unwanted or aberrant cells through apoptosis and  subsequent phagocytosis (apoptotic cell clearance) is crucial for normal  ...

The Use of Chemostats in Microbial Systems Biology

Cells regulate their rate of growth in response to signals from the external world. As the cell grows, diverse cellular processes must be coordinated including macromolecular synthesis, metabolism and ultimately, commitment to the cell division cycle. The chemostat, a method of experimentally controlling cell growth rate, provides a powerful means of systematically studying how growth rate impacts cellular processes - including gene expression and metabolism - and the regulatory networks that control the rate of cell growth. When maintained for hundreds of generations chemostats can be used to study adaptive evolution of microbes in environmental conditions that limit cell growth. We describe the principle of chemostat cultures, demonstrate their operation and provide examples of their various applications. Following a period of disuse after their introduction in the middle of the twentieth century, the convergence of genome-scale methodologies with a renewed interest in the regulation of cell growth and the molecular basis of adaptive evolution is stimulating a renaissance in the use of chemostats in biological research.

Cell growth rate is a regulated process and a primary determinant of cell physiology. Continuous culturing using chemostats enables extrinsic control of cell growth rate by nutrient limitation facilitating the study of molecular networks that control cell growth and how those networks evolve to optimize cell growth.

Der Einsatz von Chemostate in Microbial Systems Biology

Cells regulate their rate of growth in response to signals from the external world. As the cell grows, diverse cellular processes must be coordinated ...

Culturing of Human Nasal Epithelial Cells at the Air Liquid Interface

In vitro models using human primary epithelial cells are essential in understanding key functions of the respiratory epithelium in the context of microbial infections or inhaled agents. Direct comparisons of cells obtained from diseased populations allow us to characterize different phenotypes and dissect the underlying mechanisms mediating changes in epithelial cell function. Culturing epithelial cells from the human tracheobronchial region has been well documented, but is limited by the availability of human lung tissue or invasiveness associated with obtaining the bronchial brushes biopsies. Nasal epithelial cells are obtained through much less invasive superficial nasal scrape biopsies and subjects can be biopsied multiple times with no significant side effects. Additionally, the nose is the entry point to the respiratory system and therefore one of the first sites to be exposed to any kind of air-borne stressor, such as microbial agents, pollutants, or allergens. 
Briefly, nasal epithelial cells obtained from human volunteers are expanded on coated tissue culture plates, and then transferred onto cell culture inserts. Upon reaching confluency, cells continue to be cultured at the air-liquid interface (ALI), for several weeks, which creates more physiologically relevant conditions. The ALI culture condition uses defined media leading to a differentiated epithelium that exhibits morphological and functional characteristics similar to the human nasal epithelium, with both ciliated and mucus producing cells. Tissue culture inserts with differentiated nasal epithelial cells can be manipulated in a variety of ways depending on the research questions (treatment with pharmacological agents, transduction with lentiviral vectors, exposure to gases, or infection with microbial agents) and analyzed for numerous different endpoints ranging from cellular and molecular pathways, functional changes, morphology, etc. 
In vitro models of differentiated human nasal epithelial cells will enable investigators to address novel and important research questions by using organotypic experimental models that largely mimic the nasal epithelium in vivo.

Nasal epithelial cells, obtained through superficial scrape biopsy of human volunteers, are expanded and transferred onto tissue culture inserts. Upon reaching confluency, cells are grown at air liquid interface, yielding cultures of ciliated and non-ciliated cells. Differentiated nasal epithelial cell cultures provide viable experimental models for studying the respiratory mucosa.

Die Kultivierung der menschlichen Nasen Epithelzellen an der Air flüssig-Grenzfläche

In vitro models using human primary  epithelial cells are essential in understanding key functions of the respiratory  epithelium in the context of ...

Probe-based Real-time PCR Approaches for Quantitative Measurement of microRNAs

Probe-based quantitative PCR (qPCR) is a favoured method for measuring transcript abundance, since it is one of the most sensitive detection methods that provides an accurate and reproducible analysis. Probe-based chemistry offers the least background fluorescence as compared to other (dye-based) chemistries. Presently, there are several platforms available that use probe-based chemistry to quantitate transcript abundance. qPCR in a 96 well plate is the most routinely used method, however only a maximum of 96 samples or miRNAs can be tested in a single run. This is time-consuming and tedious if a large number of samples/miRNAs are to be analyzed. High-throughput probe-based platforms such as microfluidics (e.g. TaqMan Array Card) and nanofluidics arrays (e.g. OpenArray) offer ease to reproducibly and efficiently detect the abundance of multiple microRNAs in a large number of samples in a short time. Here, we demonstrate the experimental setup and protocol for miRNA quantitation from serum or plasma-EDTA samples, using probe-based chemistry and three different platforms (96 well plate, microfluidics and nanofluidics arrays) offering increasing levels of throughput.

Circulating microRNAs have recently emerged as promising and novel biomarkers for various cancers and other diseases. The goal of this article is to discuss three different probe-based real-time PCR platforms and methods that are available to quantify and determine the abundance of circulating microRNAs.

Sonde-basierte Echtzeit-PCR Ansätze zur quantitativen Messung von microRNAs

Probe-based quantitative PCR (qPCR) is a favoured method for measuring transcript abundance, since it is one of the most sensitive detection methods that ...

Use of Enzymatic Biosensors to Quantify Endogenous ATP or H2O2 in the Kidney

Enzymatic microelectrode biosensors have been widely used to measure extracellular signaling in real-time. Most of their use has been limited to brain slices and neuronal cell cultures. Recently, this technology has been applied to the whole organs. Advances in sensor design have made &#160;possible the measuring of&#160;cell signaling in blood-perfused in vivo kidneys. The present protocols list the steps needed to measure ATP and H2O2 signaling in the rat kidney interstitium. Two separate sensor designs are used for the ex vivo and in vivo protocols. Both types of sensor&#160;are coated with&#160;a thin enzymatic biolayer on top of a permselectivity layer to give fast responding, sensitive and selective biosensors. The permselectivity layer protects the signal from the interferents in biological tissue, and the enzymatic layer utilizes&#160;the sequential catalytic reaction of glycerol kinase and glycerol-3-phosphate oxidase in the presence of ATP to produce H2O2. The set of sensors used for the ex vivo studies further detected analyte by oxidation of H2O2 on a platinum/iridium (Pt-Ir) wire electrode. The sensors for the&#160;in vivo studies are instead based on the reduction of H2O2 on a mediator coated gold electrode designed for blood-perfused tissue. Final concentration changes&#160;are detected by real-time amperometry followed by calibration to known concentrations of analyte. Additionally, the specificity of the amperometric signal can be confirmed by the addition of enzymes such as catalase and apyrase that break down H2O2 and ATP correspondingly. These sensors also rely heavily on accurate calibrations before and after each experiment. The following two protocols establish the study of real-time detection of ATP and H2O2 in kidney tissues, and can be further modified to extend the described method for use in other biological preparations or whole organs.

Use of Enzymatic Biosensors to Quantify Endogenous ATP or H2O2 in the Kidney

Enzymatic microelectrode biosensors enable real-time measurements of extracellular cell signaling in biologically-relevant concentrations. The following protocols extend the applications of biosensors to the ex vivo and in vivo detection of ATP and H2O2 in the kidney.

Verwendung enzymatischer Biosensoren auf endogenes ATP oder H Quantifizieren<sub&gt; 2</sub&gt; O<sub&gt; 2</sub&gt; In der Niere

Enzymatic microelectrode biosensors have been widely used to measure extracellular signaling in real-time. Most of their use has been limited to brain ...