Ettan dige user manual

The sample labeling process was carried out according to the ettan dige user manual. Data file 28402117 aa image analysis ettan dige imager. Difference gel electrophoresis dige see glossary technology uses spectrally resolvable fluorescent protein labels cy2, cy3 and cy5 to enable multiplexing. An internal reference standard, consisting of two mixed samples used in the experiment, was labelled with cy2. Page 1 ge healthcare ettan daltsix electrophoresis system user manual page 3 12 psig. Preparation of a cell lysate compatible with cydye dige minimal labelling the example given here is that used with an escherichia coli model system see ettan dige user manual for more details. Preparation of a cell lysate compatible with cydye dige labelling the example given here is that used with an escherichia coli model system see ettan dige user guide manual for more details. Scheme showing the workflow for ettan dige system the benefits offered by ettan dige system are. Nov 28, 2019 the cydye dige fluor minimal dyes, cy3 or cy5, were used to label all treated samples and untreated control 50. Ettan ipgphor 3 isoelectric focusing unit cytiva formerly. Special tray templates that are part of the latest scanner control software and new optional gel alignment guides add higher throughput and ease of handling 2d gels run on ettan. Difference gel electrophoresis coupled with mass spectrometry dige ms is a technology used within proteomics that has demonstrated technical robustness and associated statistical confidence to enable successful identification of therapeutic targets. When you use immobiline drystrip gels for the firstdimension separation, the resultant 2d spot maps provide superior results in terms of resolution and reproducibility.

Ettan ipgphor 3 is a fully integrated isoelectric focusing ief system optimized to deliver high throughput, speed, reproducibility and high proteinloading capacity. Synchronized solutions for better 2d electrophoresis. New techniques such as 2d dige and different sample preparation methods. Decyder differential analysis user manual 18117316 edition aa. Hagnermcwhirter a, winkvist m, bourin s, marouga r. Decyder 2d software, version 6 university of hong kong.

The cell lysis solution should not contain any primary amines, such as. This process was carried out according to the ettan dige user s manual 18116440 edition aa, ge healthcare. Standard glass or plasticbacked plates can result in the generation of a high background signal. Im using lumiprobe 3dye 2d dige dyes for labeling, however after ief and 2nd dimension, i found no spots on the gel. Selective labelling of cellsurface proteins using cydye dige. Decyder 2d software user manual 28401006 ab 9 fig 11. A total of 50 g protein sample was labeled with 400 pmol of cy3 or cy5. Amersham biosciences uk ltd amersham place little chalfont buckinghamshire hp7 9na uk 80642960ac asia pacific tel. The cydye dige fluor minimal dyes, cy3 or cy5, were used to label all treated samples and untreated control 50. Five infected and noninfected samples were compared. Ettan dige fluors user manual, amersham biosciences 2004 rpk0272pl minimal dyes and. Ettan ipgphor 3 ief system is a fully integrated isoelectric focusing system optimized to deliver speed and reliability in the first dimension of 2d electrophoresis run up to 12 ipg strips 7, 11, 18, or 24 cm simultaneously using ettan ipgphor manifold, ettan ipgphor manifold light, or ceramic strip holders. Ettan ipgphor 3 isoelectric focusing system, ge healthcare. Appendix i, extracted from 2d electrophoresis, ge healthcare.

The user can create, save, and edit protocols and also pause during a run to edit a protocol in real time. Labeling was performed on ice for 30 min in the dark and the reaction was terminated by the addition of lysine to a final concentration of 1 mm. Influence of the twocomponent system saers on global gene. Resuspend cell pellets in 1 ml standard cell wash buffer in a 1. This process was carried out according to the ettan dige users manual 18116440 edition aa, ge healthcare. Ettan dige system user manual 18117317 edition ab 9 1 introduction 1. Coomassie blue, fluorescence, and labeling with ettan dige cydye fluor dyes. Experimental design for ettan dige system applications appendix i, extracted from 2d electrophoresis, ge healthcare, 2010 the differences in methodology between 2d dige and traditional 2d electrophoresis are outlined in table 41. For further information, see ettan dige system user manual see also the protein and nucleic acid sample preparation selection guide.

Gehealthcare 2006 ettan dige system user manual, isbn 18116440 ge healthcare user manuals henkel aw, bieger sc 1994 quantification of proteins dissolved in an electrophoresis sample buffer. It is a powerful tool for separating complex mixtures of. Experimental design for ettan dige system applications sigma. Ettan dige imager, including installation kit 63005642 power cable, cd with scan control software, test slides, and user manual with installation guidelines ettan dige imager cassette, 276 x 212 mm, 11002704 for dalt gel sandwiches. Comparative mitochondrial proteomic analysis of human. Dedicated userfriendly software barcode technology delivers a high degree of sample traceability. Cydye dige fluor labeling kit for scarce samples reagents for labeling protein with cydye dige fluor cy3 and cy5. Ge ettan dige imager cable kit, software version 1. Twodimensional difference gel electrophoresis dige has strengthened the 2d platform by allowing the detection and quantitation of differences between samples resolved on the same gel, or across multiple gels, when linked by an internal standard. Ettan ipgphor 3 ief system cytiva, formerly ge healthcare. Influence of the twocomponent system saers on global. Please see the ettan dige user manual for all buffer recipes and for more detailed directions the following provides only an overview. Samples were labelled with three cydye dige fluors, cy2, cy3 and cy5. A00603 ettan ipgphor ii isoelectric focusing system.

Typhoon imagers are fully optimized as part of the ettan dige system, and seamless integration with decyder differential analysis software is ensured for all models. Ettan ipgphor 3 ief system from ge healthcare used. Ettan ipgphor 3 control software generates professional graphical reports with a large user interface for programming and editing. The amersham dige protocol was followed for all steps ettan dige system user manual, ed. Carprofeninduced depletion of proton motive force reverses. Since the system variation with ettan dige system is low due to the internal standard and analysis method, biological variation will far exceed.

Amersham cydye dige fluor labeling kit for scarce samples reagents for labeling protein with cydye dige fluor cy3 and cy5 saturation dyes, before 2dimensional electrophoresis product booklet codes. For troubleshooting 2d dige results, please refer to the ettan dige user manual. Sample preparation for ettan dige system applications. To test the cell surface protein labeling protocol in an ettan dige experiment using all three dyes 6, a series of samples from serum depleted cho cells were collected and cell surface proteins labeled at different time points table 1.

Please refer to the ettan ipgphor 3 user manual for instructions on how to. Isoelectric focusing was done using an ipgphor 3 apparatus ge. The gel is cast in a low fluorescent glass cassette that is compatible with 2d dige analysis. Ge healthcare life sciences synchronized solutions for better 2d electrophoresis improved 2d dige. For creating, editing, and running protocol programs from the ettan ipgphor control panel, please refer to the ettan ipgphor 3 instrument user manual. Accurate quantitation and statistica l analysis of protein abundance changes high sensitivity and wide dynamic range 5 orders of magnitude minimization of system geltogel. Dige gel and dige buffer kit user manual available as a pdffile on the web, ettan dige system user manual 18117317 ab, and 2d electrophoresis principles and methods handbook 80642960 ac. Unlike other 2d electrophoresis methods, dige allows the option of reference to an internal standard for each spot, effectively eliminating geltogel variation and delivering highly accurate, reproducible, and dependable quantitation. Ettan dige system users manual university of alberta. Proteins were labelled according to the ettan dige minimal labelling protocol ettan dige user manual, ge healthcare.

Pdf selective labelling of cellsurface proteins using. I would much appreciate to receive a pdf of your paper. Ettan ipgphor 3 isoelectric focusing system function and description 1 ettan ipgphor 3 instrument user manual 11003458 edition ab control panel the ettan ipgphor 3 is controlled by a 7key membrane keypad. Connect only to coolant sources with regulated pressure. Ettan dige system user manual 18117317 summary proteins from prokaryotic or eukaryotic cells as discrete spots that can be quantitated. Divided into compartments with a platform in the middle, the horizontal electrophoresis systems completely submerge plated gel platforms in a buffer. Recommendations for different cell types are listed on pages 1920. Twodimensional separation of protein samples sigmaaldrich. Manual warping enables simple, interactive editing. Data file 28937442 aa 2d electrophoresis ipgbox and. The mitochondria protein lysates from two cell lines were labeled with cy2, cy3, and cy5 following the protocols described in the ettan dige user manual 18. Jul 18, 2011 2ddige, image scanning, and statistical analysis. Ge healthcare ettan ipgphor3 system used verified powerup.

Selective labelling of cellsurface proteins using cydye. Experimental design for ettan dige system applications. Ettan dige applications are described in detail in the ettan dige user manual. Twodimensional difference gel electrophoresis springerlink. Lowfluorescence glass plates must be used for gels used in ettan dige system. This user manual is broadly divided into two main parts, the reference manual chapters 111 and the tutorials chapters 17. Ettan dige imager, including installation kit 63005642 power cable, cd with scan control software, test slides, and user manual with installation guidelines ettan dige imager cassette, 276 x 212 mm, 11002704 for dalt gel sandwiches ettan dige imager cassette, 180 x 160 mm, 11002732. For detailed instructions on installation and usage of ettan ipgphor control software, and also on computer instrument connection, please refer to the ettan ipgphor software user manual. Modulation of the host cell proteome by the intracellular. I can not load it from jove web site selective labelling of cellsurface proteins using cydye dige fluor minimal dyes.

Cydye dige fluors minimal dyes for ettan dige ipmb. Ettan dige system user manual, amersham biosciences, 18116440 2002. Sample preparation for ettan dige system applications sigma. Multidimensional plasma protein separation technique for. Data visualisation flexible and powerful data display and analysis tools. Cydye dige fluor labeling kit for scarce samples plus.

Difference gel electrophoresis 2d dige analysis and is a key element in the dige system. Dige gel and dige buffer kit user manual 28946089 edition ab 5 introduction 1 1introduction dige gel is a precast polyacrylamide gel for the second dimension of 2dimensional 2d electrophoresis. If working with a sample in an unknown buffer, the sample should be precipitated and resuspended in an ettan dige systemcompatible buffer. Dige buffer kit 28937452 specially designed twobuffer system for running up to twelve dige gels simultaneously. It is recommended that new users first work thro ugh the tutorials, in order to gain a rapid understanding of the softwares ca pabilities. This page describes the differences in methodology between 2d dige and. For analytical 2ddige, denv1 and denv3 infected and noninfected protein samples were compared using the cydye dige fluors for ettan dige cy2, cy3, and cy5. Ge healthcare ettan daltsix user manual pdf download. Look this note on the user manual ge healthcare ettan dige system.

Amersham cydye dige fluor labeling kit for scarce samples. Allows accurate picking of protein spots from nonbacked 2d gels increases the versatility of ettan spot picker compatible with common staining methods same picking efficiency as for backed gels ettan spot picker nonbacked gel kit consists of a frame to. Cydye labeling and 2ddige were performed according to the ettan dige user manual amersham biosciences. Dilute anode buffer in ettan dalt electrophoresis unit for the daltsix, insert the gel cassette holder into the. For further information relating to sample preparation, please refer to the ettan dige user manual.