Protein Sample Preparation for 2-D Electrophoresis

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Overview

Efficient and reproducible sample preparation methods are key to successful 2-D electrophoresis. Sample preparation methods range from extraction with simple solubilization solutions to the use of complex mixtures of chaotropic agents, detergents, and reducing agents. Sample preparation can include enrichment strategies or separating protein mixtures into reproducible fractions to reduce their complexity.

Related Topics: Protein Extraction and Cleanup, Protein Fractionation and Depletion, 2-D Electrophoresis, and Troubleshooting 2-D Gels with 2-D Electrophoresis Doctor™.

 

Solubilization

An effective sample preparation procedure will:

  • Completely lyse the cells or tissues to release the protein content
  • Reproducibly solubilize proteins of all classes, including hydrophobic proteins
  • Prevent protein aggregation and loss of solubility during focusing
  • Prevent post-extraction chemical modification, including enzymatic or chemical degradation of the protein sample
  • Remove or thoroughly digest nucleic acids and other interfering molecules
  • Yield proteins of interest at detectable levels, which may require the removal of interfering abundant proteins or non relevant classes of proteins

 

Most protein extraction projects will require empirical determination of optimal conditions for your 2-D PAGE application. Variations in the concentrations of chaotropic agents, detergents, ampholytes, and reducing agents can dramatically affect the 2-D pattern. Find sample preparation and protein quantitation methods in the Protocols section below.

 

Related Content

 
2-D Video Tutorial
How to run a 2-D Gel from start to finish.
 
Literature
Number Description Download
2895 Protein Blotting Guide, Ver C Click to download
2651 2-D Electrophoresis Workflow How-To Guide, Rev F Click to download
Number Description Options
6200 Sample Quantitation (RC DC Protein Assay) Protocol and Compatibility Table Click to download
6221 Sample Quantitation (RC DC Protein Assay) Click to download
6220 Solubilization Click to download
 
 
LUSQH6HYP [x-forwarded-proto] = [http] [x-forwarded-port] = [80] [x-forwarded-for] = [69.166.8.43, 10.232.2.59] [seourl] = [/en-us/applications-technologies/applications-technologies/protein-sample-preparation-for-2-d-electrophoresis] [x-amzn-trace-id] = [Root=1-5c162fc2-05147582b0a8ec8009411806] [x-forwarded-server] = [lsds-prod-s.br.aws-livesite.io] [x-forwarded-host] = [www.bio-rad.com] [x-query-string] = [ID=LUSQH6HYP] [host] = [10.232.17.28:1776] [x-request-uri] = [/en-us/applications-technologies/applications-technologies/protein-sample-preparation-for-2-d-electrophoresis] [connection] = [Keep-Alive] [cache-control] = [max-age=0] [accept-encoding] = [identity] [user-agent] = [Mozilla/5.0 (compatible; Googlebot/2.1; +http://www.google.com/bot.html)] AppTech/AppTechDetails pageStyleKey internet/solutions_sub applications-technologies/applications-technologies/protein-sample-preparation-for-2-d-electrophoresis LSR LUSQH6HYP Protein Sample Preparation for 2-D Electrophoresis Protein Sample Preparation for 2-D Electrophoresis /webroot/web/html/lsr/solutions/technologies/2d_electrophoresis <p>Efficient and reproducible sample preparation methods are key to successful 2-D electrophoresis. Sample preparation methods range from extraction with simple solubilization solutions to the use of complex mixtures of chaotropic agents, detergents, and reducing agents. Sample preparation can include enrichment strategies or separating protein mixtures into reproducible fractions to reduce their complexity.</p> <p><strong>Related Topics:</strong> <a href="/evportal/destination/solutions?catID=LUSQJM7OP">Protein Extraction and Cleanup</a>, <a href="/evportal/destination/solutions?catID=LUSQKGDN">Protein Fractionation and Depletion</a>, <a href="/evportal/destination/solutions?catID=LUSQG6LPT">2-D Electrophoresis</a>, and <a href="/evportal/destination/solutions?catID=LUSQQRB9O">Troubleshooting 2-D Gels with 2-D Electrophoresis Doctor&trade;</a>.</p> <p>&nbsp;</p> <h3>Solubilization</h3> <p><img style="margin: 5px 0px 15px;" src="/evportal/framework/skins/evolution/images/1x1grey.gif" border="0" alt="" width="620" height="1" /></p> <p>An effective sample preparation procedure will:</p> <ul> <li>Completely lyse the cells or tissues to release the protein content</li> <li>Reproducibly solubilize proteins of all classes, including hydrophobic proteins</li> <li>Prevent protein aggregation and loss of solubility during focusing</li> <li>Prevent post-extraction chemical modification, including enzymatic or chemical degradation of the protein sample</li> <li>Remove or thoroughly digest nucleic acids and other interfering molecules</li> <li>Yield proteins of interest at detectable levels, which may require the removal of interfering abundant proteins or non relevant classes of proteins</li> </ul> <p>&nbsp;</p> <p>Most protein extraction projects will require empirical determination of optimal conditions for your 2-D PAGE application. Variations in the concentrations of chaotropic agents, detergents, ampholytes, and reducing agents can dramatically affect the 2-D pattern. <a href="#related_content">Find sample preparation and protein quantitation methods</a> in the Protocols section below.</p> Protocols <table id="carttablealigned" class="literature_table" style="height: auto; width: 583px;" border="0" cellspacing="0" cellpadding="0"> <tbody> <tr> <th>Number</th> <th>Description</th> <th class="options">Options</th> </tr> <tr> <td width="100">6200</td> <td width="350">Sample Quantitation (RC DC Protein Assay) Protocol and Compatibility Table</td> <td class="pdf"><a class="pdf" href="/webroot/web/pdf/lsr/literature/Bulletin_6200.pdf" target="_blank"><span>Click to download</span></a></td> </tr> <tr> <td>6221</td> <td>Sample Quantitation (RC DC Protein Assay)</td> <td class="pdf"><a class="pdf" href="/webroot/web/pdf/lsr/literature/Bulletin_6221.pdf" target="_blank"><span>Click to download</span></a></td> </tr> <tr> <td>6220</td> <td>Solubilization</td> <td class="pdf"><a class="pdf" href="/webroot/web/pdf/lsr/literature/Bulletin_6220.pdf" target="_blank"><span>Click to download</span></a></td> </tr> </tbody> </table> <div class="videowrap vwrap_last"> <div class="videoImg"><a title="2-D Video Tutorial" onclick="javascript:openAjaxOverlay('/webroot/web/html/lsr/products/electrophoresis/product_overlay/global/2-d_tutorial_video.html');" href="javascript:void(0);"> <img style="border:none" src="/webroot/web/images/lsr/support/tutorials/global/2d_tutorial.jpg" alt="" /></a></div> <div class="videoDesc"><a title="2-D Video Tutorial" onclick="javascript:openAjaxOverlay('/webroot/web/html/lsr/products/electrophoresis/product_overlay/global/2-d_tutorial_video.html');" href="javascript:void(0);">2-D Video Tutorial</a><br /> How to run a 2-D Gel from start to finish.</div> <div class="clear">&nbsp;</div> </div> 2895 2651 Life Science Research/Products/Protein Sample Preparation/Protein Extraction/MicroRotofor Cell Lysis Kits ->MT::5e97c5be-3707-4c08-ae55-25e883f2ab99##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/Protein Sample Preparation/Protein Extraction/ReadyPrep Protein Extraction Kit (Total Protein) ->MT::749eaa83-8fb4-4854-bc8a-84ab01195fb9##Life Science Research/Products/Protein Sample Preparation/Protein Sample Cleanup/ReadyPrep 2-D Cleanup Kit ->MT::bbf3a0be-188f-45c7-b81e-e9706ec77c02##Life Science Research/Products/Protein Sample Preparation/Protein Sample Cleanup/ReadyPrep Reduction-Alkylation Kit ->MT::9c6fff0c-677e-440d-9795-49d17aa08aae##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 Research/Solutions/Applications/2-D Electrophoresis and Analysis ->MTS::LUSQF04EH## Life Science Research/Solutions/Technologies/Protein Electrophoresis/Protein Detection and Analysis/Protein Staining ->MTS::LUSPMPE8Z##Life Science Research/Solutions/Technologies/2-D Electrophoresis/Imaging and Analysis ->MTS::LUSQON470##Life Science Research/Solutions/Technologies/Protein Electrophoresis ->MTS::LUSOVO47B##Life Science Research/Solutions/Technologies/Protein Electrophoresis/Protein Sample Preparation ->MTS::LUSP108WI## Karen Moss 12/13/11 04:35 PM 12/13/21 04:47 PM 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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