Bu çalışma, Avrupa Birliği IST Programı (IST-2002-507.464 Projesi &quot;- Fobiler &#39;Tedavi için Sanal Gerçeklik Akıllı Çoklu-sensör Giyilebilir Sistem INTREPID&quot;) tarafından desteklenmiştir.

Yaygın Anksiyete Bozukluğu tedavisi için klinik protokolü (YAB), terapistin ofis içinde ve dışında aynı anda programlanmıştır . Gevşeme programı gerginliğe ve çeşitli kas gruplarını rahatlatıcı, zıt duyumları dikkate alırken ve &quot;Güdümlü Görüntü&quot; programı içeren &quot;İlerici Kas gevşemesi Eğitimi&quot; (PMRT), Jacobsen tarafından tanımlanan klasik prosedür, bir kombinasyonudur. Eğitim görüntüleri rahatlatıcı bir sanal ortamda görsel ipucu tarafından değiştirilir. Terapistin OFİS İÇ. Hastalar dört hafta (haftada 2 seans) dağıtılan terapist ile 8 eğitim oturumları, katılmak. Her seans hastanın anksiyete ölçü veren bir klinik değerlendirme ile başlar ve durur. Terapistin ofisinde oturumları Planı: <ol><li> Hastanın durumuna ilk klinik değerlendirme; </li><li> Hasta / fizyolojik parametreleri (deri iletkenliği, kalp atım hızı, solunum) kayıt biyosensörler bağlıdır. Bu parametrelerin bir temel önlemi 3 dakika kayıtlı olduğu; </li><li> Hasta bir pc ile bağlantılı bir baş-üstü ekran giyer ve bir joystick kolları; </li><li> Okyanusa bakan güzel bir tropik ada: Hasta, sanal ortamda keşfetmek başlar. Hasta, terapist tarafından kaydedilen anlatı, tekne ile adaya ulaşır ve onu keşfetmek. Bir yaya yolu takiben kılavuzları ona / onu adaya farklı paneller farklı hedef alanlara yönleri gösterir başlangıç ​​noktası, hastanın gelir. Bu alanların her birinde rahatlatıcı bir egzersiz verilir; sanal ortamda bazı unsurları doğrudan onun tarafından güncellenmiştir / onun fizyolojik parametreler kaydedildi ise bu eğitim sırasında, sesli rehber tarafından verilen endikasyon, hasta onu / kendini rahatlamak için çalışır gerçek zamanlı olarak. Bu nedenle, hasta / aktivasyonu (geleneksel biofeedback teknikleri gibi) onu hemen bir geri bildirim alır, ama o / o immersively keşfetmek olduğunu sanal ortamda verilen ekstra değer. Hedef alanlara bağlı: <ul><li> Campfire: fizyolojik parametrelere, yangın bir azalma hastanın fizyolojik aktivasyon sonuçları azaltma sönene kadar böylece yangın yoğunluğu, kontrolü; </li><li> Dalgalar: okyanus tamamen sakin bir hale gelinceye kadar fizyolojik parametrelere dalgaları kontrolü; </li><li> Şelale: su düşmeye durana kadar fizyolojik bir uyarılma azalma akım şiddetinin azalmasına karşılık gelir. </li><li> Gazebo: diğerlerinden farklı, bu egzersiz, kelime ya da ona / onu kişisel stresli olaylarla ilgili görüntüleri seçebilirsiniz hasta tarafından özelleştirilebilir. Bu teknik bir stres aşılama programı olarak hizmet verir ve hastanın seçilen öğenin boyut-solmaya yüzen azalma yaşayabilirsiniz onu / kendini dinlenmek için onun yeteneklerini / bağlı olarak; </li></ul></li><li> Sonra sanal gerçeklik oturumu tamamlandı, fizyolojik parametreler, geri kalanı durumda tekrar kaydedilir; </li><li> Hastanın durumuna Final klinik değerlendirme. </li></ol> DIŞ terapistin OFİS: eğitim etkinliğini artırmak ve rahatlama etkisi artırmak amacıyla, hastaların terapistin ofis dışında Evde Senaryo yaşayarak rahatlama teknikleri uygulama olacaktır. Bu tedavi sırasında bir mobil cihazda yaşanan aynı sanal ortam sunarak gerçekleştirilmektedir. Mobil cihaz, kalp hızı ve deri iletkenlik Biyosensör bir dizi Bluetooth üzerinden bağlanır. Bu cihaz, yeterince küçük ve özerk (bkz. Şekil 1) hasta tarafından ele alınması, kullanımı kolay. Hasta, terapist tarafından sağlanan tedavi planı, tedavi süresinin tamamını kapsayan (bir ay) için günde en az bir kez gevşeme yetenekleri eğitmek için gereklidir. Temsilcisi Sonuçlar Biofeedback ve sanal gerçeklik de gevşeme etkili olduğu bilinmektedir, çünkü ana beklenen sonuç bu iki yöntem bir arada bireysel olarak kullanılan iki teknik daha yaygın anksiyete bozukluğu etkilenen hastaların, anksiyete azaltmada daha etkili olmasıdır. <img src="/files/ftp_upload/1554/Fig1.jpg" alt="Şekil 1" /> Şekil 1: GSR / HR biyosensörler modülü.

<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.

Bizim hipotezi, sanal gerçeklik ve sadece sanal gerçeklik ile doğrudan karşılaştırıldığında, sanal ortamda değiştirir biofeedback kombinasyonu, daha iyi ve daha hızlı bir rahatlama öğrenmeye yol açacak. Eğer öyleyse, bu yeni araç, anksiyete bozuklukları tedavi etmek için oluşturulan uygulamalı ve diğer anksiyete ile ilgili patolojilerde bile test edilebilir.

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

Yaygın anksiyete bozukluğu (YAB), günlük yaşam aktiviteleri ile müdahale sürekli ve belirsiz anksiyete ile karakterize bir psikiyatrik bozukluk. Genel popülasyonda yüksek prevalansı ve neden olan ciddi sınırlamalar, onu tedavi etmek için yeni ve etkili stratejiler bulmak için gerekliliğine işaret etmektedir. Bilişsel-davranışçı tedaviler ile birlikte, dinlenme GAD tedavisi için yararlı bir yaklaşımı temsil eder, ama öğrenilmesi gereken zor sınırlaması vardır. INTREPID projesi, anksiyete ile ilgili bozuklukların tedavisinde ve anksiyete ile ilgili belirtileri azaltmada, klinik etkinliğini test etmek için yeni bir enstrüman uygulanması amaçlanmıştır. İlki, doğrudan ikinci bir çıkış olarak değiştirilmiş, böylece bu yaklaşımın yenilik, sanal gerçeklik ve biofeedback birleşimidir. Bu şekilde, hasta, gerçek zamanlı olarak VR ortamında bazı özellikleri değişiklik ile onun reaksiyonlar farkında yapılır. Onun başarısını ölçmek için zihinsel egzersizler hastanın bu fizyolojik parametrelerini kontrol etmeyi öğrenir ve sanal ortamda verilen geribildirim kullanarak yapabiliyor. PDA veya akıllı telefonları gibi taşınabilir cihazlar, tamamlayıcı kullanımı, hastanın, ayrı ayrı ve özerk, evde terapistin ofisinde deneyimli aynı egzersizleri yapmak için izin verir. Amaç, bu nedenle hastanın kendi semptomları ile başa çıkmak için yeteneğini arttırmak, gerçek bir yaşam bağlamında öğrenilen protokol çapa. Beklenen sonucu geleneksel klinik protokoller ile karşılaştırıldığında, böylece tedavinin bir artış etkinliğini rahatlama teknikleri öğrenme daha iyi ve daha hızlı, ve.

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

Anksiyete Bozuklukları (INTREPID) Tedavi için Sanal Gerçeklik Akıllı Çoklu sensör Sistem Projesi anksiyete bozukluklarının tedavisi için çoklu sensör, bağlam-duyarlı bir sanal gerçeklik sistemi geliştirilmesi hedeflenmektedir.

Klinik Sanal Gerçeklik Biofeedback kullanımı: INTREPID Projesi

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

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Research

JoVE Journal

Biology

15.0K Görüntüleme.  Istituto Auxologico Italiano. Anksiyete Bozuklukları (INTREPID) Tedavi için Sanal Gerçeklik Akıllı Çoklu sensör Sistem Projesi anksiyete bozukluklarının tedavisi için çoklu sensör, bağlam-duyarlı bir sanal gerçeklik sistemi geliştirilmesi hedeflenmektedir.

Video: 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.

Rotorod Rat Yürüyüş Kalitatif Analiz için bir yöntem olarak kullanımı

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

Biyoloji

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.

Biofeedback Kullanımı Psikofizyolojik Stres Değerlendirme

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.

Proteaz Aktivitesi belirleme Proteaz Floresan Algılama Kit kullanın

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.

Kendini yenileme LIF Yokluğunda Mesc Destek SC1 (Pluripotin) kullanımı

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.

Saflaştırılması M. magneticum Deformasyon AMB-1 Magnetosome İlişkili 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.

Apoptotik hücresi Temizliği görüntülenmesi sırasında Canlı<em&gt; Drosophila</em&gt; Embriyo

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.

Mikrobiyal Systems Biyoloji Chemostats Kullanımı

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.

Hava Sıvı Arayüz İnsan Burun Epitel Hücrelerinin ekimi

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.

Probe-tabanlı Real-time PCR microRNA Kantitatif Ölçüm Yaklaşımları

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.

Enzimatik Biyosensörlerin Kullanımı Endojen ATP veya H belirlememize<sub&gt; 2</sub&gt; O<sub&gt; 2</sub&gt; Böbrek

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