2-D Electrophoresis

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Overview

en-us LUSQG6LPT 2-D Electrophoresis 2-D Electrophoresis /webroot/web/html/lsr/solutions/technologies/2d_electrophoresis /webroot/web/images/lsr/solutions/technologies/2-D_electrophoresis/2-D_electrophoresis/technology_detail/2-D-electrophoresis-feature.jpg /webroot/web/images/lsr/solutions/technologies/2-D_electrophoresis/technology_detail/cat_2dt1_2d_electro_icon.jpg Picture of 2-D electrophoresis gel <script type="text/javascript"><!-- if ($.browser.msie && $.browser.version < 8) {$("div.methodboxmiddle ul.rightarrowsearch1").css({"margin-left":"-5px"});} $('head').append('<meta name="DCSext.at_banner" content="2-D Electrophoresis" /><meta name="DCSext.at_banner_e" content="v" />'); // --></script> <p>Proteomics, the analysis of the complete complement of proteins in a cell, tissue, or organism (the proteome), involves the detection of the presence or absence of proteins and the direct measurement of relative protein abundances. One of the greatest challenges of proteome analysis is the reproducible fractionation of complex protein mixtures while retaining the qualitative and quantitative relationships among component proteins. Currently, two-dimensional polyacrylamide gel electrophoresis (2-D PAGE), which is capable of resolving thousands of proteins in a single run, is the primary tool of proteomics research. This section describes the various steps of a typical 2-D electrophoresis workflow, including</p> <br /> <ul> <li>Protein sample preparation</li> <li>First-dimension electrophoresis</li> <li>Second-dimension electrophoresis</li> <li>Gel staining and protein visualization</li> <li>Gel imaging and protein analysis</li> <li>Protein identification</li> </ul> <p>A special section, the <a href="/evportal/destination/solutions?catID=LUSQQRB9O">2-D Doctor&trade;</a>, is a practical resource for 2-D gel troubleshooting.</p> <p>Proteome analysis is used to determine which proteins in a cell, tissue, or organism are affected by changes in conditions such as disease states or developmental stages. Protein profiles in different states of a cell or an organism can be compared to identify proteins that are qualitatively and quantitatively affected by the condition of interest. Such profiling requires superior protein separation resolution and high-throughput technologies to address the potentially large numbers of proteins.</p> <p>2-D electrophoresis can be used to resolve complex mixtures of thousands of proteins. In the first dimension, proteins are separated based on differences in isoelectric point (pI). In the second dimension, they are separated according to molecular weight. Following separation, 2-D electrophoresis gels are stained for protein visualization and analysis. In combination with computer-assisted image evaluation systems for comprehensive qualitative and quantitative examination of proteomes, this electrophoresis technique allows cataloging of proteins and comparison of data among groups of researchers.</p> 2-D Electrophoresis Workflow <p><img style="display: block; margin: 0 auto;" src="/webroot/web/images/lsr/solutions/technologies/2-D_electrophoresis/2-D_electrophoresis/technology_detail/2-d-electrophoresis-workflow-chart.jpg" border="0" alt="Diagram of Bio-Rad&rsquo;s 2-D electrophoresis workflow" /></p> <p class="caption"><strong>The 2-D electrophoresis workflow.</strong></p> <p>The general workflow of a 2-D gel-based proteomics experiment is outlined below, and some of the factors affecting the way the experiment is performed are discussed. <a href="/evportal/destination/commerce/product_detail?catID=MGSKID15">Bio-Rad&rsquo;s 2-D Electrophoresis Workflow system</a> provides a comprehensive set of product solutions and educational resources to help you achieve better 2-D results.</p> <ul> <li><strong><a href="/evportal/destination/solutions?catID=LUSQH6HYP">Protein Sample Preparation</a></strong><br /> The method of sample preparation depends on the aim of the research and is key to the success of the experiment. Factors such as the solubilities, sizes, charges, and isoelectric points (pI) of the proteins of interest are considerations for sample preparation. Sample preparation is also important for reducing the complexity of a protein mixture. A protein fraction to be separated by 2-D PAGE must be prepared in a denaturing buffer of low ionic strength that maintains the native charges of the proteins and keeps them soluble.Bio-Rad offers a variety of kits ranging from the basic <a href="/evportal/destination/commerce/product_detail?catID=6a7bf285-fd23-43c4-bdd8-c4f8012d4aef">ReadyPrep&trade; 2-D starter kit</a> to the <a href="/evportal/destination/commerce/product_detail?catID=bbf3a0be-188f-45c7-b81e-e9706ec77c02">ReadyPrep 2-D cleanup kit</a> and <a href="/evportal/destination/commerce/product_detail?catID=1dd94f06-7658-4ab4-b844-e29a1342a214">ProteoMiner&trade; protein enrichment kit</a> for cleanup and enrichment of low-abundance proteins in complex protein samples</li> <li><strong><a href="/evportal/destination/solutions?catID=LUSQLK2B7">First-Dimension Separation (Isoelectric Focusing)</a></strong><strong></strong><br /> Proteins are first separated on the basis of pI, the pH at which a protein carries no net charge and thus will not migrate in an electrical field. The technique is called isoelectric focusing (IEF). For 2-D PAGE, IEF is best performed in an immobilized pH gradient (IPG) gel strip, which can subsequently be directly applied onto a PAGE gel for second-dimension separation. Bio-Rad&rsquo;s <a href="/evportal/destination/commerce/product_detail?catID=56994cbe-ea1b-48d8-b7e9-bc17726c0d70">ReadyStrip&trade; IPG strips</a> offer many pH gradient and strip length options. The <a href="/evportal/destination/commerce/product_detail?catID=92682b19-051c-464f-8a73-97cd256b225f">PROTEAN<sup>&reg;</sup> i12&trade; IEF</a> system provides the flexibility to run multiple pH gradients, protocols, and samples simultaneously with built-in independent lane controls</li> <li><strong><a href="/evportal/destination/solutions?catID=LUSQMI97Q">Second-Dimension Separation</a></strong><strong></strong><br /> In the next step of 2-D electrophoresis, an anionic surfactant such as sodium dodecyl sulfate (SDS) is typically added to impart a uniform negative charge to the proteins per unit mass and so insure uniform separation based on their molecular weights. The choice of SDS-PAGE second-dimension gel properties such as polyacrylamide percentage and gradient depends on the molecular weight (MW) range of the proteins to be separated and the size of the IPG strip used in the first dimension. The ability to run many gels at the same time under the same conditions is important for the purpose of gel-to-gel comparison. Bio-Rad&rsquo;s Criterion&trade; system includes <a href="/evportal/destination/commerce/product_detail?catID=LAAU8515">Criterion precast gels</a> in many gradient options and provides the ability to run up to 12 gels at once in the <a href="/en-us/product/second-dimension-midi-format-electrophoresis-systems/criterion-dodeca-cell">Dodeca&trade; cell</a>. <a href="/evportal/destination/commerce/product_detail?catID=0c7564b9-7eec-48d8-b071-068a67e04ce4">Precision Plus Protein&trade; standard plugs</a> allow loading of unstained molecular weight standards on vertical 2&ndash;D gels with no reference well</li> <li><strong><a href="/evportal/destination/solutions?catID=LUSQN83Q3">2-D Gel Staining</a></strong><br /> To visualize proteins in 2-D electrophoresis gels, the proteins must be stained or labeled. The choice of staining method is determined by several factors including desired sensitivity, linear range, ease of use, expense, and the type of imaging equipment available. Bio-Rad&rsquo;s <a href="/evportal/destination/commerce/product_detail?catID=cb886820-1f6e-447f-8ef6-772fd81c29e3">Oriole&trade; fluorescent gel stain</a> is a one-step stain for quickly visualizing 2-D gels for image analysis and spot cutting. The separated proteins can also be detected and quantitated by <a href="/evportal/destination/solutions?catID=LUSPPAKG4">western blotting</a> after transfer to a membrane support</li> <li><strong><a href="/evportal/destination/solutions?catID=LUSQON470">2-D Gel Imaging</a></strong><br /> The ability to collect data in digital form is a major factor in making 2-D electrophoresis a practical means for collecting proteomics information. Digital gel imaging allows unprejudiced comparison of gels, the transfer of information among research groups, and cataloging of immense amounts of data. Many types of imaging devices interface with software designed specifically to collect, interpret, and compare proteomics data. Bio-Rad's <a href="/en-us/product/chemidoc-mp-imaging-system">ChemiDoc&trade; MP</a> and <a href="/en-us/product/chemidoc-imaging-systems/chemidoc-xrs-system">ChemiDoc XRS</a> imaging systems feature multiplex fluorescence, chemiluminescence, and colorimetric detection and can accommodate a variety of sample types</li> <li><strong><a href="/evportal/destination/solutions?catID=LUSQON470">2-D Gel Image Analysis</a></strong><br /> Bio-Rad's <a href="/evportal/destination/commerce/product_detail?catID=966deb78-2656-437f-b7a4-ab0a9bd45c8d">PDQuest<sup>&trade;</sup> software</a> and similar image analysis software packages compare gel images, annotate protein spots, and catalog data. These software packages facilitate proteomics experiments by enabling the comparison of large 2-D electrophoresis data sets. </li> <li><strong><a href="/evportal/destination/solutions?catID=LUSQPTOZR">Protein Identification</a></strong><br /> Once proteins of interest are selected by differential analysis or other criteria, the proteins can be excised from gels and identified. The ability to precisely determine MW by mass spectrometry and to search databases for peptide mass matches has made high-throughput protein identification possible</li> </ul> <div class="bannerAT"> <div class="bannerText ddpcrText"> <h2 class="banner_header">Simplify your 2-D electrophoresis workflow.</h2> <p style="width: 420px;">Browse a wide selection of ReadyStrip&trade; Immobilized pH Gradient (IPG) Strips for first-dimension IEF &mdash; wide and narrow ranges.</p> <a class="linkgeneration" style="padding-right:15px;" href="/_locale/product/readystrip-ipg-strips?at_banner=2-D Electrophoresis&amp;at_banner_rev=2-D Electrophoresis&amp;at_banner_e=c">Learn more &raquo;</a></div> <img src="/webroot/web/images/lsr/global/english/solutions/readystrip-ipg-strips.jpg" alt="ReadyStrip&trade; IPG Strips" width="615" height="120" /></div> <div class="top"><a href="#helptop">Back to Top</a></div> <script src="/webroot/web/js/countrySpecific-min.js" type="text/javascript"></script> <script type="text/javascript"><!-- $(document).ready(function(){ setSterlingUrlsToHtmlHrefVariables(); $('a.linkgeneration').each(function(){ $(this).attr('href', $(this).attr('href').replace('_locale', languageCode + '-' + countryCode).replace('_verticalUrl', currentVerticalUrlTitle).replace('_defaultVerticalUrl', defaultVerticalUrlTitle).replace('_feedbackCMSID',feedbackCMSID)); }); }); // --></script> 6478 /templatedata/internet/documentation/data/LSR/Literature/6478_1401994020.xml 2-D Electrophoresis Workflow Brochure, Rev B 6478 H /webroot/web/pdf/lsr/literature/Bulletin_6478.pdf Literature PDF Brochures_and_Specifications 2-D Electrophoresis Workflow Brochure, Rev B No 2-D Electrophoresis Workflow Brochure, Rev B 6478 6478, 2D electrophoresis, 2-D, ReadyPrep, PROTEAN i12, ProteoMiner, Bio-Lyte, BioLyte, ReadyStrip, Criterion Dodeca, ChemiDoc, Flamingo, Silver Stain Plus, EXQuest, TGX Stain-Free, Oriole, IEF separation, isoelectric focusing 6493 /templatedata/internet/documentation/data/LSR/Literature/6493_1401994287.xml Bio-Rad’s Host Cell Protein (HCP) Workflow Quick Guide 6493 H /webroot/web/pdf/lsr/literature/Bulletin_6493.pdf Literature PDF Manuals_and_Quick_Guides Bio-Rad’s Host Cell Protein (HCP) Workflow Quick Guide No Bio-Rad’s Host Cell Protein (HCP) Workflow Quick Guide, Rev C 6493 6493, hcp, host cell, protein, two-dimensional, 2-d, 2de, electrophoresis, western blotting, readyprep, protean, readystrip, criterion tgx, precision plus, oriole, trans-blot turbo, trans blot, transblot, chemidoc, chemidoc touch, clarity, pdquest, ief separation, isoelectric focusing, compliance, 161-0378, 161-0496, 163-2000, 163-2002, 163-2003, 163-2001, 163-2004, 163-2005, 163-2014, 163-2016, 163-2017, 163-2015, 163-2018, 163-2019, 163-2007, 163-2009, 163-2010, 163-2008, 163-2011, 163-2012, 163-2032, 163-2033, 163-2035, 163-2034, 163-2036, 163-2037, 163-2042, 163-2043, 163-2045, 163-2044, 163-2046, 163-2047, 164-6000, 165-6001, 170-3125, 170-4155, 170-4275, 170-3127, 170-5060, 170-5061, 170-8280, 170-8370, 170-9630, 567-1071, 567-1081, 567-1091, 567-1101, 567-1111, 567-1121, 1610378, 1610496, 1632000, 1632002, 1632003, 1632001, 1632004, 1632005, 1632014, 1632016, 1632017, 1632015, 1632018, 1632019, 1632007, 1632009, 1632010, 1632008, 1632011, 1632012, 1632032, 1632033, 1632035, 1632034, 1632036, 1632037, 1632042, 1632043, 1632045, 1632044, 1632046, 1632047, 1646000, 1656001, 1703125, 1704155, 1704275, 1703127, 1705060, 1705061, 1708280, 1708370, 1709630, 5671071, 5671081, 5671091, 5671101, 5671111, 5671121 6040 /templatedata/internet/documentation/data/LSR/Literature/6040.xml Electrophoresis Guide, Interactive PDF, Rev C 6040 /webroot/web/pdf/lsr/literature/Bulletin_6040.pdf Literature PDF Manuals_and_Quick_Guides /webroot/web/images/general/icons/icon_pdf.gif No Electrophoresis Guide, Interactive PDF, Rev C 6040 Bulletin 2895, 6040, overview and principle, theory and product selection, one dimensional (1-D) protein separation, blue native polyacrylamide, SDS-PAGE, sodium dodecyl sulfate, sodium dodecyl sulphate, polymerization, precast, handcast resolving, stacking single-percentage gel, Laemmli (Tris-HCl), Bis-Tris, Tris-acetate, Tris-Tricine premade running and sample buffers, formulations/components, reagents, salts, isoelectric focusing (IEF), zymogram, gel casting chambers, gradient formers, sample preparation protocols, cell lysis, disruption, homogenization, protein solubilization, human suspension/monolayer cultured cells, mammalian tissue, plant leaves, microbial cultures, chromatography protein fractions, reducing, chaotropic agents, detergents, sample loading and quantitation spectrophotometric assays, recombinant and natural protein standards, prestained or unstained molecular weight markers or ladders, selecting power supplies, joule heating, separations under constant voltage, current, power, detection and analysis, total and specific protein staining, Coomassie Blue stain, fluorescent gel and silver stains, stainers, imagers, imaging systems and software, protein molecular weight (size) estimation, quantification, downstream applications, western blotting, gel drying, electroelution, spot excision (cutting), troubleshooting tips 2895 Protein Blotting Guide, Ver C 2895 /webroot/web/pdf/lsr/literature/Bulletin_2895.pdf Literature PDF Other /webroot/web/images/general/icons/icon_pdf.gif Protein Blotting Guide No Protein Blotting Guide, Ver C 2895 Bulletin 2895, horseradish peroxidase (HRP), enzyme-antibody complex, alkaline phosphatase (AP), nitroblue tetrazolium (NBT) tablets, BCIP, secondary antibody probe, amido black, streptavidin-biotin, anionic dye, total protein blotting stain, nitrocellulose, supported nitrocellulose or polyvinylidene difluoride (PVDF) or low fluorescence LF PVDF membranes, antibody, immunoglobulin (IgG), antigen, assay, avidin, biotin, background, non-specific noise, Bjerrum Schafer-Nielsen buffer, blocking reagent, Western blotting, BLOTTO, chemiluminescence, colloidal gold, color development reagent, colorimetric detection, chemiluminescent detection, enzyme-antibody conjugate, Coomassie Blue stain, diaminobenzidine (DAB), dot blot, Dunn buffer, electrophoretic blotting, foam pads, filter paper, gelatin, high-intensity transfer, immunoassay, immunoblotting, immunodetection, ligand, membrane, membrane/filter paper sandwiches, microfiltration blotting, multiplexing, multiscreen apparatus, native PAGE, polyacrylamide gel electrophoresis, NHS-biotin, non-enzymatic probe, nonenzymatic probe, non-fat dry milk, non-specific binding, phycobiliprotein, power supply, primary antibody, prestained standards, molecular weight markers or ladders, Protein A, Protein G, rapid semi-dry blotting, SDS-PAGE, sodium dodecyl sulfate, sodium dodecyl sulphate, signal-to-noise ratio, signal to noise ratio, Stain-free technology, CCD imager, charge-coupled device, StrepTactin, Strep-tag sequence, substrate, super cooling coil, tank blotting, Towbin buffer, Tween 20 2651 2-D Electrophoresis Workflow How-To Guide, Rev F 2651 /webroot/web/pdf/lsr/literature/Bulletin_2651.pdf Literature PDF Manuals_and_Quick_Guides /webroot/web/images/general/icons/icon_pdf.gif 2-D Electrophoresis Workflow How-To Guide No 2-D Electrophoresis Workflow How-To Guide, Rev F 2651 proteomics, bulletin 2651, two dimensional electrophoresis, LIT2651, sample preparation, 2d, analytical, proteomeworks, prefractionation, 2-d manual, 2-d guide, 2-d electrophoresis workflow, 2-d methods and product, 2d manual, 2d guide, 2d electrophoresis workflow, 2d methods and product 6441 /templatedata/internet/documentation/data/LSR/Literature/6441.xml 2-D Analysis of Leaf Protein Samples Treated with ProteoMiner Beads Under Denaturing and Nondenaturing Conditions Poster, Rev A 6441 /webroot/web/pdf/lsr/literature/Bulletin_6441.pdf Literature PDF Scientific_Posters /webroot/web/images/general/icons/icon_pdf.gif 2-D Analysis of Leaf Protein Samples Treated with ProteoMiner Beads Under Denaturing and Nondenaturing Conditions No 2-D Analysis of Leaf Protein Samples Treated with ProteoMiner Beads Under Denaturing and Nondenaturing Conditions Poster, Rev A 6441 2-D Analysis of Leaf Protein Samples Treated with ProteoMiner Beads Under Denaturing and Nondenaturing Conditions Life Science Research/Products/Electrophoresis and Blotting/Protein Electrophoresis and Blotting/2D-Electrophoresis/2-D Buffers and Reagents/ReadyPrep 2-D starter kit ->MT::6a7bf285-fd23-43c4-bdd8-c4f8012d4aef##Life Science Research/Products/Electrophoresis and Blotting/Protein Electrophoresis and Blotting/2D-Electrophoresis/1st Dimension- Isoelectric Focusing (IEF)/PROTEAN IEF Cell ->MT::92682b19-051c-464f-8a73-97cd256b225f##Life Science Research/Products/Electrophoresis and Blotting/Protein Electrophoresis and Blotting/2D-Electrophoresis/1st Dimension- Isoelectric Focusing (IEF)/ReadyStrip IPG Strips ->MT::56994cbe-ea1b-48d8-b7e9-bc17726c0d70##Life Science Research/Products/Electrophoresis and Blotting/Protein Electrophoresis and Blotting/Mini Format 1D-Electrophoresis Systems/Mini-PROTEAN Tetra Cell Systems ->MT::5cf78e19-7ed5-4373-a988-3e62456a488e##Life Science Education/Products/Equipment and Supplies/Reagents/Protein Stains ->MT::ef400f0d-45fa-4b0d-b410-1b7593add5fa## Life Science Research/Solutions/Applications/2-D Electrophoresis and Analysis ->MTS::LUSQF04EH## Life Science Research/Solutions/Technologies/Protein Electrophoresis ->MTS::LUSOVO47B##Life Science Research/Solutions/Technologies/Protein Electrophoresis/Protein Detection and Analysis/Protein Staining ->MTS::LUSPMPE8Z##Life Science Research/Solutions/Technologies/Imaging and Analysis ->MTS::LUSQC6MNI## Karen Moss 2D Protein Electrophoresis <p>Useful information from sample preparation to imaging of 2-D gels, along with protocols, video tutorials, demonstrations, and troubleshooting tips.</p> 12/06/11 10:06 AM 12/06/21 10:42 AM AE,AI,AL,AM,AR,AT,AU,AZ,BA,BD,BE,BF,BG,BH,BN,BO,BR,BW,CA,CH,CL,CM,CN,CO,CR,CY,CZ,DE,DK,DO,DZ,EC,EE,EG,EH,ER,ES,ET,FI,FM,FO,FR,GA,GE,GF,GH,GP,GR,GT,GU,HK,HN,HR,HT,HU,ID,IE,IL,IN,IS,IT,JM,JO,JP,KE,KH,KR,KW,KZ,LB,LI,LK,LT,LU,LV,MA,MD,MG,MK,ML,MO,MQ,MS,MT,MU,MX,MY,NG,NI,NL,NO,NP,NZ,OM,PA,PE,PF,PG,PH,PK,PL,PR,PS,PT,PW,PY,QA,RO,RS,RU,SA,SB,SE,SG,SI,SK,SN,ST,SV,TG,TH,TN,TO,TR,TT,TW,TZ,UA,UG,UK,US,UY,UZ,VA,VE,VU,XK,YE,ZA en LSR /LSR/Technologies/2-D_Electrophoresis N 0 Troubleshooting 2-D Electrophoresis Gels with 2-D Doctor&trade; /en-us/applications-technologies/applications-technologies/troubleshooting-2-d-electrophoresis-gels-with-2-d-doctor?ID=LUSQQRB9O Protein Sample Preparation for 2-D Electrophoresis /en-us/applications-technologies/applications-technologies/protein-sample-preparation-for-2-d-electrophoresis?ID=LUSQH6HYP Isoelectric Focusing in 2D Electrophoresis /en-us/applications-technologies/applications-technologies/isoelectric-focusing-2d-electrophoresis?ID=LUSQLK2B7 Second-Dimension Separation /en-us/applications-technologies/applications-technologies/second-dimension-separation?ID=LUSQMI97Q Staining and Visualization of Proteins After 2-D Electrophoresis /en-us/applications-technologies/applications-technologies/staining-visualization-proteins-after-2-d-electrophoresis?ID=LUSQN83Q3 Imaging and Analysis of 2-D Electrophoresis Gels /en-us/applications-technologies/applications-technologies/imaging-analysis-2-d-electrophoresis-gels?ID=LUSQON470 2D Protein Spot Excision & Detection /en-us/applications-technologies/applications-technologies/2d-protein-spot-excision-detection?ID=LUSQPTOZR Technologies /en-us/applications-technologies/applications-technologies/2-d-electrophoresis?ID=LUSLZJC4S

Proteomics, the analysis of the complete complement of proteins in a cell, tissue, or organism (the proteome), involves the detection of the presence or absence of proteins and the direct measurement of relative protein abundances. One of the greatest challenges of proteome analysis is the reproducible fractionation of complex protein mixtures while retaining the qualitative and quantitative relationships among component proteins. Currently, two-dimensional polyacrylamide gel electrophoresis (2-D PAGE), which is capable of resolving thousands of proteins in a single run, is the primary tool of proteomics research. This section describes the various steps of a typical 2-D electrophoresis workflow, including


  • Protein sample preparation
  • First-dimension electrophoresis
  • Second-dimension electrophoresis
  • Gel staining and protein visualization
  • Gel imaging and protein analysis
  • Protein identification

A special section, the 2-D Doctor™, is a practical resource for 2-D gel troubleshooting.

Proteome analysis is used to determine which proteins in a cell, tissue, or organism are affected by changes in conditions such as disease states or developmental stages. Protein profiles in different states of a cell or an organism can be compared to identify proteins that are qualitatively and quantitatively affected by the condition of interest. Such profiling requires superior protein separation resolution and high-throughput technologies to address the potentially large numbers of proteins.

2-D electrophoresis can be used to resolve complex mixtures of thousands of proteins. In the first dimension, proteins are separated based on differences in isoelectric point (pI). In the second dimension, they are separated according to molecular weight. Following separation, 2-D electrophoresis gels are stained for protein visualization and analysis. In combination with computer-assisted image evaluation systems for comprehensive qualitative and quantitative examination of proteomes, this electrophoresis technique allows cataloging of proteins and comparison of data among groups of researchers.

 
 
Page Contents
 
 
 

2-D Electrophoresis Workflow

Diagram of Bio-Rad’s 2-D electrophoresis workflow

The 2-D electrophoresis workflow.

The general workflow of a 2-D gel-based proteomics experiment is outlined below, and some of the factors affecting the way the experiment is performed are discussed. Bio-Rad’s 2-D Electrophoresis Workflow system provides a comprehensive set of product solutions and educational resources to help you achieve better 2-D results.

  • Protein Sample Preparation
    The method of sample preparation depends on the aim of the research and is key to the success of the experiment. Factors such as the solubilities, sizes, charges, and isoelectric points (pI) of the proteins of interest are considerations for sample preparation. Sample preparation is also important for reducing the complexity of a protein mixture. A protein fraction to be separated by 2-D PAGE must be prepared in a denaturing buffer of low ionic strength that maintains the native charges of the proteins and keeps them soluble.Bio-Rad offers a variety of kits ranging from the basic ReadyPrep™ 2-D starter kit to the ReadyPrep 2-D cleanup kit and ProteoMiner™ protein enrichment kit for cleanup and enrichment of low-abundance proteins in complex protein samples
  • First-Dimension Separation (Isoelectric Focusing)
    Proteins are first separated on the basis of pI, the pH at which a protein carries no net charge and thus will not migrate in an electrical field. The technique is called isoelectric focusing (IEF). For 2-D PAGE, IEF is best performed in an immobilized pH gradient (IPG) gel strip, which can subsequently be directly applied onto a PAGE gel for second-dimension separation. Bio-Rad’s ReadyStrip™ IPG strips offer many pH gradient and strip length options. The PROTEAN® i12™ IEF system provides the flexibility to run multiple pH gradients, protocols, and samples simultaneously with built-in independent lane controls
  • Second-Dimension Separation
    In the next step of 2-D electrophoresis, an anionic surfactant such as sodium dodecyl sulfate (SDS) is typically added to impart a uniform negative charge to the proteins per unit mass and so insure uniform separation based on their molecular weights. The choice of SDS-PAGE second-dimension gel properties such as polyacrylamide percentage and gradient depends on the molecular weight (MW) range of the proteins to be separated and the size of the IPG strip used in the first dimension. The ability to run many gels at the same time under the same conditions is important for the purpose of gel-to-gel comparison. Bio-Rad’s Criterion™ system includes Criterion precast gels in many gradient options and provides the ability to run up to 12 gels at once in the Dodeca™ cell. Precision Plus Protein™ standard plugs allow loading of unstained molecular weight standards on vertical 2–D gels with no reference well
  • 2-D Gel Staining
    To visualize proteins in 2-D electrophoresis gels, the proteins must be stained or labeled. The choice of staining method is determined by several factors including desired sensitivity, linear range, ease of use, expense, and the type of imaging equipment available. Bio-Rad’s Oriole™ fluorescent gel stain is a one-step stain for quickly visualizing 2-D gels for image analysis and spot cutting. The separated proteins can also be detected and quantitated by western blotting after transfer to a membrane support
  • 2-D Gel Imaging
    The ability to collect data in digital form is a major factor in making 2-D electrophoresis a practical means for collecting proteomics information. Digital gel imaging allows unprejudiced comparison of gels, the transfer of information among research groups, and cataloging of immense amounts of data. Many types of imaging devices interface with software designed specifically to collect, interpret, and compare proteomics data. Bio-Rad's ChemiDoc™ MP and ChemiDoc XRS imaging systems feature multiplex fluorescence, chemiluminescence, and colorimetric detection and can accommodate a variety of sample types
  • 2-D Gel Image Analysis
    Bio-Rad's PDQuest software and similar image analysis software packages compare gel images, annotate protein spots, and catalog data. These software packages facilitate proteomics experiments by enabling the comparison of large 2-D electrophoresis data sets.
  • Protein Identification
    Once proteins of interest are selected by differential analysis or other criteria, the proteins can be excised from gels and identified. The ability to precisely determine MW by mass spectrometry and to search databases for peptide mass matches has made high-throughput protein identification possible

Browse a wide selection of ReadyStrip™ Immobilized pH Gradient (IPG) Strips for first-dimension IEF — wide and narrow ranges.

Learn more »
ReadyStrip™ IPG Strips
 

Related Content

 
Literature
Number Description Download
6478 2-D Electrophoresis Workflow Brochure, Rev B Click to download
6493 Bio-Rad’s Host Cell Protein (HCP) Workflow Quick Guide Click to download
6040 Electrophoresis Guide, Interactive PDF, Rev C Click to download
2895 Protein Blotting Guide, Ver C Click to download
2651 2-D Electrophoresis Workflow How-To Guide, Rev F Click to download
6441 2-D Analysis of Leaf Protein Samples Treated with ProteoMiner Beads Under Denaturing and Nondenaturing Conditions Poster, Rev A Click to download
 
 
LUSQG6LPT [x-forwarded-proto] = [http] [accept-language] = [en] [x-forwarded-port] = [80] [x-forwarded-for] = [144.76.137.254, 10.232.3.215] [accept] = [text/html,text/plain,text/xml,text/*,application/xml,application/xhtml+xml,application/rss+xml,application/atom+xml,application/rdf+xml,application/php,application/x-php,application/x-httpd-php] [seourl] = [/en-us/applications-technologies/experion-automated-electrophoresis-system] [x-amzn-trace-id] = [Root=1-5b755699-b99b1835721b0742e793a76a] [x-forwarded-server] = [lsds-prod-s.br.aws-livesite.io] [x-forwarded-host] = [lsds-prod-s.br.aws-livesite.io] [x-query-string] = [ID=LUSQG6LPT] [host] = [10.232.0.21:1776] [x-request-uri] = [/en-us/applications-technologies/experion-automated-electrophoresis-system] [connection] = [Keep-Alive] [accept-encoding] = [gzip] [user-agent] = [Mozilla/5.0 (compatible; MJ12bot/v1.4.8; http://mj12bot.com/)] AppTech/AppTechDetails pageStyleKey internet/solutions_sub applications-technologies/experion-automated-electrophoresis-system LSR LUSQG6LPT 2-D Electrophoresis 2-D Electrophoresis /webroot/web/html/lsr/solutions/technologies/2d_electrophoresis /webroot/web/images/lsr/solutions/technologies/2-D_electrophoresis/2-D_electrophoresis/technology_detail/2-D-electrophoresis-feature.jpg /webroot/web/images/lsr/solutions/technologies/2-D_electrophoresis/technology_detail/cat_2dt1_2d_electro_icon.jpg Picture of 2-D electrophoresis gel <script type="text/javascript"><!-- if ($.browser.msie && $.browser.version < 8) {$("div.methodboxmiddle ul.rightarrowsearch1").css({"margin-left":"-5px"});} $('head').append('<meta name="DCSext.at_banner" content="2-D Electrophoresis" /><meta name="DCSext.at_banner_e" content="v" />'); // --></script> <p>Proteomics, the analysis of the complete complement of proteins in a cell, tissue, or organism (the proteome), involves the detection of the presence or absence of proteins and the direct measurement of relative protein abundances. One of the greatest challenges of proteome analysis is the reproducible fractionation of complex protein mixtures while retaining the qualitative and quantitative relationships among component proteins. Currently, two-dimensional polyacrylamide gel electrophoresis (2-D PAGE), which is capable of resolving thousands of proteins in a single run, is the primary tool of proteomics research. This section describes the various steps of a typical 2-D electrophoresis workflow, including</p> <br /> <ul> <li>Protein sample preparation</li> <li>First-dimension electrophoresis</li> <li>Second-dimension electrophoresis</li> <li>Gel staining and protein visualization</li> <li>Gel imaging and protein analysis</li> <li>Protein identification</li> </ul> <p>A special section, the <a href="/evportal/destination/solutions?catID=LUSQQRB9O">2-D Doctor&trade;</a>, is a practical resource for 2-D gel troubleshooting.</p> <p>Proteome analysis is used to determine which proteins in a cell, tissue, or organism are affected by changes in conditions such as disease states or developmental stages. Protein profiles in different states of a cell or an organism can be compared to identify proteins that are qualitatively and quantitatively affected by the condition of interest. Such profiling requires superior protein separation resolution and high-throughput technologies to address the potentially large numbers of proteins.</p> <p>2-D electrophoresis can be used to resolve complex mixtures of thousands of proteins. In the first dimension, proteins are separated based on differences in isoelectric point (pI). In the second dimension, they are separated according to molecular weight. Following separation, 2-D electrophoresis gels are stained for protein visualization and analysis. In combination with computer-assisted image evaluation systems for comprehensive qualitative and quantitative examination of proteomes, this electrophoresis technique allows cataloging of proteins and comparison of data among groups of researchers.</p> 2-D Electrophoresis Workflow <p><img style="display: block; margin: 0 auto;" src="/webroot/web/images/lsr/solutions/technologies/2-D_electrophoresis/2-D_electrophoresis/technology_detail/2-d-electrophoresis-workflow-chart.jpg" border="0" alt="Diagram of Bio-Rad&rsquo;s 2-D electrophoresis workflow" /></p> <p class="caption"><strong>The 2-D electrophoresis workflow.</strong></p> <p>The general workflow of a 2-D gel-based proteomics experiment is outlined below, and some of the factors affecting the way the experiment is performed are discussed. <a href="/evportal/destination/commerce/product_detail?catID=MGSKID15">Bio-Rad&rsquo;s 2-D Electrophoresis Workflow system</a> provides a comprehensive set of product solutions and educational resources to help you achieve better 2-D results.</p> <ul> <li><strong><a href="/evportal/destination/solutions?catID=LUSQH6HYP">Protein Sample Preparation</a></strong><br /> The method of sample preparation depends on the aim of the research and is key to the success of the experiment. Factors such as the solubilities, sizes, charges, and isoelectric points (pI) of the proteins of interest are considerations for sample preparation. Sample preparation is also important for reducing the complexity of a protein mixture. A protein fraction to be separated by 2-D PAGE must be prepared in a denaturing buffer of low ionic strength that maintains the native charges of the proteins and keeps them soluble.Bio-Rad offers a variety of kits ranging from the basic <a href="/evportal/destination/commerce/product_detail?catID=6a7bf285-fd23-43c4-bdd8-c4f8012d4aef">ReadyPrep&trade; 2-D starter kit</a> to the <a href="/evportal/destination/commerce/product_detail?catID=bbf3a0be-188f-45c7-b81e-e9706ec77c02">ReadyPrep 2-D cleanup kit</a> and <a href="/evportal/destination/commerce/product_detail?catID=1dd94f06-7658-4ab4-b844-e29a1342a214">ProteoMiner&trade; protein enrichment kit</a> for cleanup and enrichment of low-abundance proteins in complex protein samples</li> <li><strong><a href="/evportal/destination/solutions?catID=LUSQLK2B7">First-Dimension Separation (Isoelectric Focusing)</a></strong><strong></strong><br /> Proteins are first separated on the basis of pI, the pH at which a protein carries no net charge and thus will not migrate in an electrical field. The technique is called isoelectric focusing (IEF). For 2-D PAGE, IEF is best performed in an immobilized pH gradient (IPG) gel strip, which can subsequently be directly applied onto a PAGE gel for second-dimension separation. Bio-Rad&rsquo;s <a href="/evportal/destination/commerce/product_detail?catID=56994cbe-ea1b-48d8-b7e9-bc17726c0d70">ReadyStrip&trade; IPG strips</a> offer many pH gradient and strip length options. The <a href="/evportal/destination/commerce/product_detail?catID=92682b19-051c-464f-8a73-97cd256b225f">PROTEAN<sup>&reg;</sup> i12&trade; IEF</a> system provides the flexibility to run multiple pH gradients, protocols, and samples simultaneously with built-in independent lane controls</li> <li><strong><a href="/evportal/destination/solutions?catID=LUSQMI97Q">Second-Dimension Separation</a></strong><strong></strong><br /> In the next step of 2-D electrophoresis, an anionic surfactant such as sodium dodecyl sulfate (SDS) is typically added to impart a uniform negative charge to the proteins per unit mass and so insure uniform separation based on their molecular weights. The choice of SDS-PAGE second-dimension gel properties such as polyacrylamide percentage and gradient depends on the molecular weight (MW) range of the proteins to be separated and the size of the IPG strip used in the first dimension. The ability to run many gels at the same time under the same conditions is important for the purpose of gel-to-gel comparison. Bio-Rad&rsquo;s Criterion&trade; system includes <a href="/evportal/destination/commerce/product_detail?catID=LAAU8515">Criterion precast gels</a> in many gradient options and provides the ability to run up to 12 gels at once in the <a href="/en-us/product/second-dimension-midi-format-electrophoresis-systems/criterion-dodeca-cell">Dodeca&trade; cell</a>. <a href="/evportal/destination/commerce/product_detail?catID=0c7564b9-7eec-48d8-b071-068a67e04ce4">Precision Plus Protein&trade; standard plugs</a> allow loading of unstained molecular weight standards on vertical 2&ndash;D gels with no reference well</li> <li><strong><a href="/evportal/destination/solutions?catID=LUSQN83Q3">2-D Gel Staining</a></strong><br /> To visualize proteins in 2-D electrophoresis gels, the proteins must be stained or labeled. The choice of staining method is determined by several factors including desired sensitivity, linear range, ease of use, expense, and the type of imaging equipment available. Bio-Rad&rsquo;s <a href="/evportal/destination/commerce/product_detail?catID=cb886820-1f6e-447f-8ef6-772fd81c29e3">Oriole&trade; fluorescent gel stain</a> is a one-step stain for quickly visualizing 2-D gels for image analysis and spot cutting. The separated proteins can also be detected and quantitated by <a href="/evportal/destination/solutions?catID=LUSPPAKG4">western blotting</a> after transfer to a membrane support</li> <li><strong><a href="/evportal/destination/solutions?catID=LUSQON470">2-D Gel Imaging</a></strong><br /> The ability to collect data in digital form is a major factor in making 2-D electrophoresis a practical means for collecting proteomics information. Digital gel imaging allows unprejudiced comparison of gels, the transfer of information among research groups, and cataloging of immense amounts of data. Many types of imaging devices interface with software designed specifically to collect, interpret, and compare proteomics data. Bio-Rad's <a href="/en-us/product/chemidoc-mp-imaging-system">ChemiDoc&trade; MP</a> and <a href="/en-us/product/chemidoc-imaging-systems/chemidoc-xrs-system">ChemiDoc XRS</a> imaging systems feature multiplex fluorescence, chemiluminescence, and colorimetric detection and can accommodate a variety of sample types</li> <li><strong><a href="/evportal/destination/solutions?catID=LUSQON470">2-D Gel Image Analysis</a></strong><br /> Bio-Rad's <a href="/evportal/destination/commerce/product_detail?catID=966deb78-2656-437f-b7a4-ab0a9bd45c8d">PDQuest<sup>&trade;</sup> software</a> and similar image analysis software packages compare gel images, annotate protein spots, and catalog data. These software packages facilitate proteomics experiments by enabling the comparison of large 2-D electrophoresis data sets. </li> <li><strong><a href="/evportal/destination/solutions?catID=LUSQPTOZR">Protein Identification</a></strong><br /> Once proteins of interest are selected by differential analysis or other criteria, the proteins can be excised from gels and identified. The ability to precisely determine MW by mass spectrometry and to search databases for peptide mass matches has made high-throughput protein identification possible</li> </ul> <div class="bannerAT"> <div class="bannerText ddpcrText"> <h2 class="banner_header">Simplify your 2-D electrophoresis workflow.</h2> <p style="width: 420px;">Browse a wide selection of ReadyStrip&trade; Immobilized pH Gradient (IPG) Strips for first-dimension IEF &mdash; wide and narrow ranges.</p> <a class="linkgeneration" style="padding-right:15px;" href="/_locale/product/readystrip-ipg-strips?at_banner=2-D Electrophoresis&amp;at_banner_rev=2-D Electrophoresis&amp;at_banner_e=c">Learn more &raquo;</a></div> <img src="/webroot/web/images/lsr/global/english/solutions/readystrip-ipg-strips.jpg" alt="ReadyStrip&trade; IPG Strips" width="615" height="120" /></div> <div class="top"><a href="#helptop">Back to Top</a></div> <script src="/webroot/web/js/countrySpecific-min.js" type="text/javascript"></script> <script type="text/javascript"><!-- $(document).ready(function(){ setSterlingUrlsToHtmlHrefVariables(); $('a.linkgeneration').each(function(){ $(this).attr('href', $(this).attr('href').replace('_locale', languageCode + '-' + countryCode).replace('_verticalUrl', currentVerticalUrlTitle).replace('_defaultVerticalUrl', defaultVerticalUrlTitle).replace('_feedbackCMSID',feedbackCMSID)); }); }); // --></script> 6478 /templatedata/internet/documentation/data/LSR/Literature/6478_1401994020.xml 6493 /templatedata/internet/documentation/data/LSR/Literature/6493_1401994287.xml 6040 /templatedata/internet/documentation/data/LSR/Literature/6040.xml 2895 2651 6441 /templatedata/internet/documentation/data/LSR/Literature/6441.xml Life Science Research/Products/Electrophoresis and Blotting/Protein Electrophoresis and Blotting/2D-Electrophoresis/2-D Buffers and Reagents/ReadyPrep 2-D starter kit ->MT::6a7bf285-fd23-43c4-bdd8-c4f8012d4aef##Life Science Research/Products/Electrophoresis and Blotting/Protein Electrophoresis and Blotting/2D-Electrophoresis/1st Dimension- Isoelectric Focusing (IEF)/PROTEAN IEF Cell ->MT::92682b19-051c-464f-8a73-97cd256b225f##Life Science Research/Products/Electrophoresis and Blotting/Protein Electrophoresis and Blotting/2D-Electrophoresis/1st Dimension- Isoelectric Focusing (IEF)/ReadyStrip IPG Strips ->MT::56994cbe-ea1b-48d8-b7e9-bc17726c0d70##Life Science Research/Products/Electrophoresis and Blotting/Protein Electrophoresis and Blotting/Mini Format 1D-Electrophoresis Systems/Mini-PROTEAN Tetra Cell Systems ->MT::5cf78e19-7ed5-4373-a988-3e62456a488e##Life Science Education/Products/Equipment and Supplies/Reagents/Protein Stains ->MT::ef400f0d-45fa-4b0d-b410-1b7593add5fa## Life Science Research/Solutions/Applications/2-D Electrophoresis and Analysis ->MTS::LUSQF04EH## Life Science Research/Solutions/Technologies/Protein Electrophoresis ->MTS::LUSOVO47B##Life Science Research/Solutions/Technologies/Protein Electrophoresis/Protein Detection and Analysis/Protein Staining ->MTS::LUSPMPE8Z##Life Science Research/Solutions/Technologies/Imaging and Analysis ->MTS::LUSQC6MNI## Karen Moss 2D Protein Electrophoresis <p>Useful information from sample preparation to imaging of 2-D gels, along with protocols, video tutorials, demonstrations, and troubleshooting tips.</p> 12/06/11 10:06 AM 12/06/21 10:42 AM AE,AI,AL,AM,AR,AT,AU,AZ,BA,BD,BE,BF,BG,BH,BN,BO,BR,BW,CA,CH,CL,CM,CN,CO,CR,CY,CZ,DE,DK,DO,DZ,EC,EE,EG,EH,ER,ES,ET,FI,FM,FO,FR,GA,GE,GF,GH,GP,GR,GT,GU,HK,HN,HR,HT,HU,ID,IE,IL,IN,IS,IT,JM,JO,JP,KE,KH,KR,KW,KZ,LB,LI,LK,LT,LU,LV,MA,MD,MG,MK,ML,MO,MQ,MS,MT,MU,MX,MY,NG,NI,NL,NO,NP,NZ,OM,PA,PE,PF,PG,PH,PK,PL,PR,PS,PT,PW,PY,QA,RO,RS,RU,SA,SB,SE,SG,SI,SK,SN,ST,SV,TG,TH,TN,TO,TR,TT,TW,TZ,UA,UG,UK,US,UY,UZ,VA,VE,VU,XK,YE,ZA en LSR /LSR/Technologies/2-D_Electrophoresis N 0
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