Here, we describe a detailed protocol on how dc metabolism can be studied after tlr stimulation using the 96well format extracellular flux xf e 96 analyzer from seahorse bioscience, a machine that allows one to simultaneously assess rates of oxidative phosphorylation and glycolysis in realtime, in live cells and in a highthroughput manner. Seahorse bioscience liaises with volpi for xfe extracellular. Glycolysis is determined through measurements of the extracellular acidification rate ecar of the surrounding media, which is predominately from the excretion of lactic acid per unit time. Seahorse bioscience introduces xfe the worlds most advanced. Technical specifications for the seahorse xfp extracellular flux analyzer we are seeking to procure a seahorse xfp extracellular flux analyzer, which measures bioenergetics of live mammalian and bacterial cells. The seahorse bioscience xf e extracellular flux analyzer measures the two major energy pathways of the cell mitochondrial respiration and glycolysis in a microplate, in realtime. A dedicated room air incubator is kept adjacent to the analyzer for premeasurement equilibration. Ultrasonicator ultrasonic homogenizer, model 3000, biologics, inc. Seahorse eastern virginia medical school evms, norfolk. Xfe wave software, designed for the xfe extracellular flux analyzer. View online or download seahorse xf96 installation and operation manual. Used with the seahorse xfp extracellular flux analyzer, the xf cell energy phenotype test kit measures the utilization of each major energy pathway of a cell mitochondrial respiration and. Seahorse xfe96 extracellular flux analyzer the seahorse enables quantitation of metabolomics of live cells in real time with stimulation with up to 4 drugs.
Jul 01, 2014 pancreatic cancer cells with different genetic backgrounds were analysed in realtime using a seahorse extracellular flux xf 96well analyzer. Seahorse bioscience the seahorse xf prep station automatically prepares your culture plates for assay in the xf analyzer improving reproducibility, and simplifying your workflow. Introduction to the xf software and graphic user interface 33. Xfe24 extracellular flux analyzer the xf e 24 and optional plate preparation station require two people to lift and handle safely.
The measurement of cellular bioenergetics on live cells enables timeresolved analysis and testing of multiple conditions per assay well. Experiments are designed using the xf reader software on a touch screen, and can be analyzed using excel spreadsheets. Advances in the quantification of mitochondrial function. Seahorse xfe 96 is the worlds most advanced cell metabolic analyzer and employs fiber optics based excitation and measurement technology from volpi, an expert in the development and. Seahorse extracellular flux analyzers agilent institute. Seahorse core university of nebraska medical center. The seahorse extracellular flux xf24 analyzer provides a comprehensive assessment of the relative state of glycolytic and aerobic metabolism in live cells making this method instrumental in.
Oxygen consumption rate of caenorhabditis elegans as a high. Seahorse bioscience introduces xfe the worlds most. Collection of cell analysis resources agilent community. Extracellular flux an overview sciencedirect topics.
Seahorse xf e 24 extracellular flux analyzer based analysis of cellular respiration in caenorhabditis elegans anthony l. The xfp extracellular flux analyzer is built on innovative and proven xf technology for live cellbased assays of energy metabolism. Seahorse xf96 installation and operation manual pdf download. Xfp analyzer overview the seahorse bioscience xfp analyzer measures the rate of change of dissolved oxygen and ph in the media immediately surrounding living cells cultured in a miniplate.
Seahorse consumable items can be ordered through the dna core. The following items are included in the xfe24 instrument and controller boxes instrumentcontroller quantity. A novel highthroughput assay for respiration in isolated brain. Analysis of lymphocyte mitochondrial function has been facilitated by introduction of 96well format extracellular flux xf96 analyzers, but the technology. Apr 04, 2019 xfe extracellular flux analyzer user guide version 2. The systems can be used for screening and doseresponse studies. The system should be equipped with latest version of hardware and software. Sunnybrook research institute sri is home to the worlds most advanced metabolic data analyzer. Therefore, with the development of stateoftheart, highthroughput respirometers, such as the seahorse xf e 96 extracellular flux analyzer, the relevance and applicability of c. Compact and easy to use, the xfp analyzer is designed to perform standard xf assays, and is ideal for pairwise comparisons. At sparc, we use an xfe96 extracellular flux analyzer from agilent seahorse. The seahorse xf prep station is a compact, benchtop, fluid management system that prepares your culture plates for assay with the seahorse xfe or xf extracellular flux analyzers, saving valuable lab time, improving cvs, and increasing the sensitivity of your ecar measurements. Xf core facilities facilities that provide access to agilent seahorse xf technology, including advice in experimental design, data analysis, basic training, and bioinformatics on a cost recovery basis. Nicholas school of the environment, duke university, durham, north carolina.
Cytomine single cell analysis system, exoview tetraspanin kits, flow. This product eliminates the need for radioactivity in fatty acid oxidation. A complete platform that includes hardware, software, consumables and reagents, the seahorse xfp analyzer is designed to. Seahorse xfe96 extracellular flux analyzer dartlab. Training is compulsory for firsttime users, and can be arranged upon request. Seahorse xfe 24 extracellular flux analyzerbased analysis of cellular respiration in caenorhabditis elegans. The 24well seahorse system provides researchers the opportunity to perform mediumthroughput, realtime measurement of mitochondrial function and cellular bioenergetics in an expedited manner compared to traditional techniques. Seahorse xf24 extracellular flux analyzer department of. This instrument measures oxygen consumption rate ocr and extracellular acidification rate ecar of living cells simultaneously in a 24 well format.
The seahorse core facility operates in the department of pharmacology and experimental neuroscience on the campus of unmc. Seahorse xf e 24 extracellular flux analyzerbased analysis of cellular respiration in caenorhabditis elegans anthony l. Seahorse xfe 24 extracellular flux analyzerbased analysis. Xf analyzers perform compound addition and mixing, labelfree analytical detection, and automatic measurement of ocr. Xfe24 extracellular flux assay kit 100777004, agilent technologies. The seahorse extracellular flux xf analyzer seahorse bioscience is a powerful tool for measuring glycolysis and oxidative phosphorylation through oxygen consumption simultaneously in the same cells. Extracellular flux analyzer and prep station brand. Seahorse xfp extracellular flux analyzer the role of metabolism in cellular and physiological processes from cardiovascular function to immune response is well established, with many diseases now linked to metabolic dysfunction or reprogramming. The xf e 24 analyzer determines in vitro oxygen consumption rate. Consumables include flux paks, microplates, and stress kits. Seahorse xf cell energy phenotype test kit receives top 10. Seahorse xfe24 extracellular flux analyzerbased analysis of. When prompted, transfer the sensor cartridge and utility plate to the xf e extracellular flux analyzer, click im ready, and wait for the completion of calibration see note 33. View and download seahorse xf96 installation and operation manual online.
The seahorse xf24 extracellular analyzer is located in room d2383 on the second floor of the thomas building. Xfe extracellular flux analyzers simultaneously interrogate the two major energy producing pathways of the cell mitochondrial respiration and glycolysis in a microplate, in realtime. The seahorse bioscience xf e 96 extracellular flux analyzer simultaneously measures oxygen consumption rate and extracellular acidification rate of living cells cultured in disposable 96well microplates it is the first such system to join the roster of scientific equipment. The seahorse bioscience xf e 96 extracellular flux analyzer simultaneously measures oxygen consumption rate and extracellular acidification rate of living cells cultured in disposable 96well microplates it is the first such system to join the roster of scientific. Seahorse xf analyzers measure oxygen consumption rate ocr and extracellular acidification rate ecar of live cells in a multiwell plate, interrogating key cellular functions such as mitochondrial respiration and glycolysis.
Oxygen consumption rate of caenorhabditis elegans as a. The seahorse bioscience xfe analyzer measures the rate of change of dissolved. Agilent seahorse xf palmitatebsa fao substrate quickstart guide. Each person should firmly grasp the base of the unit at opposite ends to one another. Xf analyzer grant expeditor package simplify your grant application process with the agilent seahorse xf analyzer grant expeditor package. Bioenergetics, seahorse xf, mitochondrial metabolism, glycolysis. The measurement of cellular bioenergetics on live cells enables timeresolved. The two major cellular energyyielding pathways, mitochondrial respiration and glycolysis, can be measured simultaneously and in realtime using the seahorse xfe96 instrument. Can i operate the agilent seahorse xf analyzer myself. Measurement of mitochondrial respiration and glycolysis allows energetic phenotyping of cells, in order to assess functional changes. Measuring energy pathways with the seahorse xf analyzer. Please refer to sop for costsharing department of biochemistry users for costsharing of seahorse xf e 24 extracellular flux analyzer contacts.
Seahorse bioscience sells instruments and consumables. The first published seahorse data was reported in a 2006 nature article. The xf96 extracellular flux analyzer was developed to provide higher throughput analysis for customers desiring to study mitochondrial function and dysfunction, obesity, diabetes, cancer, ageing disorders, cardiac disease and drug toxicity. Changes in these two energy pathways affect the immediate extracellular environment, and the resulting changes in o2 concentration oxygen consumption or ocr, and ph extracellular. The seahorse xf flux analyzer equipment is available for recharge use at the uc davis. Agilent seahorse xf analyzer is a sensitive, robust, highthroughput. Seahorse xf24 cellular bioenergetics analysis shared. Xfe extracellular flux analyzer user guide version 2. Required equipment, consumables, and software the following equipment, consumables, and software are required in addition to the materials specified in the icell cardiomyocytes users guide. The facility provides expertise and training in use of the agilent seahorse extracellular flux analyzer, which has become the gold standard platform for metabolic assays. Numerous studies show that mitochondrial energy generation determines the effectiveness of immune responses. Seahorse bioscience xf e 96 extracellular flux analyzer. The xf96 and xfe96 extracellular flux analyzer are 96 well tissue culture microplate systems allowing for a larger number of assays. Measuring energy pathways with the seahorse xf analyzer 19 november 2014 watch this video to learn how the seahorse xf analyzer simultaneously measures mitochondrial respiration and glycoloysis via extracellular flux, eliminating the need to use labels that may alter cell behavior.
Xf prep station automated assay plate preparation youtube. Pdf comprehensive method for culturing embryonic dorsal. Changes in the extracellular media are caused by the consumption or production of analytes by the cells. Does anyone know how to solve a problem with a negative. This fast and sensitive measurement of cellular bioenergetics is label free, enabling timeresolved analysis and. Seahorse xf e 24 extracellular flux analyzerbased analysis.
Seahorse bioscience announces the xf96 extracellular flux. Seahorse xf24 extracellular flux analyzer the seahorse xf24 is a revolutionary instrument for studies of cellular energy metabolism. The xf instruments analyze mitochondrial respiration and glycolysis in live cells. Tool kit seahorse bioscience xfe96 extracellular flux. Seahorse wave desktop software is the assay design and data analysis software for all agilent seahorse xf analyzers. Seahorse xfe analyzers from agilent cell analysis selectscience. The xf extracellular flux analyzer is the bestselling instrument. Seahorse xfe24 extracellular flux analyzerbased analysis. Xfe96 flux packs are sold individually at the cic when available. Advances in the quantification of mitochondrial function in. The seahorse analyzer is housed in the internal medicine department and is available to all researchers through user fees. This includes a disposable sensor cartridge, cell culture microplate, calibration plate, and xf calibrant. Agilent seahorse xf cell energy phenotype test kit. The power of xf technology for every lab seahorse xfp extracellular flux analyzer.
The seahorse xf e 24 extracellular flux analyzer simultaneously interrogates the two major energy producing pathways of the cell mitochondrial respiration and glycolysis in a microplate, in realtime. Nov 22, 2010 seahorse bioscience the seahorse xf prep station automatically prepares your culture plates for assay in the xf analyzer improving reproducibility, and simplifying your workflow. Seahorse xf96 installation and operation manual pdf. Seahorse bioscience xf96 extracellular flux analyzer w xf. Pancreatic cancer cells with different genetic backgrounds were analysed in realtime using a seahorse extracellular flux xf 96well analyzer. Both cellular ocr, resulting from oxidative phosphorylation, and ecar, associated with glycolytic metabolism, were simultaneously monitored in six pancreatic cancer cell lines figure 2a.
Please refer to sop for costsharing department of biochemistry users for costsharing of seahorse xf e 24 extracellular flux analyzer. Bmsb j520 reservations calendar extracellular flux or xf assays measure oxygen, total acid, and carbon dioxide in cell media and provide accuracies comparable to. Seahorse bioscience xf96 extracellular flux analyzer product details the xf96 extracellular flux analyzer is a fullyintegrated instrument that simultaneously measures the two major energy yielding pathways aerobic respiration and glycolysis in a convenient, microplate format. Seahorse xfe96 analyzers measure ocr and ecar of live cells in a 96well plate. The xf palmitatebsa fao reagent is a solution absorbed by cells. The agilent cell analysis software portfolio provides easytolearn, flexible tools solutions for designing assays, instrument operation, analyzing data, and. The seahorse bioscience xf e 96 extracellular flux analyzer simultaneously measures oxygen consumption rate and extracellular acidification rate of living cells cultured in disposable 96well microplates.
The xf96 and xf e 96 are validated for assays performed in hypoxia chambers and may be used with. Furthermore, changes in mitochondrial function may regulate lymphocyte function in inflammatory diseases like type 2 diabetes. Based analysis of cellular respiration in caenorhabditis elegans. Here, using the seahorse xf e 24 extracellular flux analyzer seahorse bioscience, massachusetts, usa we describe how to measure the fundamental parameters of the electron transport chain etc. Seahorse bioscience xf96 extracellular flux analyzer. I want to do assays using a seahorse flux analyzer 96well, mostly mitostress test with primary macrophages and i wonder, how i can normalize the results for the cell number per well after the.
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