Attomole protein analysis by CIEF with LIF detection
We have coupled CIEF with an LIF detector that is based on a post-column sheath flow cuvette. We employed Chromeo P503 as a fluorogenic reagent to label proteins before analysis. This reagent reacts with the ε-amine of lysine residues, preserving the cationic nature of the residue; labeled proteins...
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| Veröffentlicht in: | Electrophoresis 2009, Vol.30 (2), p.297-302 |
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| creator | Ramsay, Lauren M Dickerson, Jane A Dovichi, Norman J |
| description | We have coupled CIEF with an LIF detector that is based on a post-column sheath flow cuvette. We employed Chromeo P503 as a fluorogenic reagent to label proteins before analysis. This reagent reacts with the ε-amine of lysine residues, preserving the cationic nature of the residue; labeled proteins generate extremely sharp peaks in CIEF. A set of four standard proteins generated a linear relationship between migration time and pI. A protein homogenate prepared from a Barrett's esophagus cell line resolved over 100 components in a 40 min separation. Detection limits for Chromeo P503-labeled β-lactoglobulin were 5 amol injected into the capillary. Fluorescent impurities present in the ampholytes generated a large background signal that degraded the detection limit by four orders of magnitude compared with other forms of capillary electrophoresis with this detector. |
| doi_str_mv | 10.1002/elps.200800498 |
| format | Article |
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We employed Chromeo P503 as a fluorogenic reagent to label proteins before analysis. This reagent reacts with the ε-amine of lysine residues, preserving the cationic nature of the residue; labeled proteins generate extremely sharp peaks in CIEF. A set of four standard proteins generated a linear relationship between migration time and pI. A protein homogenate prepared from a Barrett's esophagus cell line resolved over 100 components in a 40 min separation. Detection limits for Chromeo P503-labeled β-lactoglobulin were 5 amol injected into the capillary. Fluorescent impurities present in the ampholytes generated a large background signal that degraded the detection limit by four orders of magnitude compared with other forms of capillary electrophoresis with this detector.</description><identifier>ISSN: 0173-0835</identifier><identifier>EISSN: 1522-2683</identifier><identifier>DOI: 10.1002/elps.200800498</identifier><identifier>PMID: 19204946</identifier><language>eng</language><publisher>Weinheim: Wiley-VCH Verlag</publisher><subject>Attomole protein analysis ; Calibration ; Cell Line ; Cells - chemistry ; CIEF ; Electrophoresis, Capillary - methods ; Fluorescent Dyes - chemistry ; LIF ; Proteins - analysis ; Sensitivity and Specificity ; Spectrometry, Fluorescence - methods ; Time Factors</subject><ispartof>Electrophoresis, 2009, Vol.30 (2), p.297-302</ispartof><rights>Copyright © 2009 WILEY‐VCH Verlag GmbH & Co. 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We employed Chromeo P503 as a fluorogenic reagent to label proteins before analysis. This reagent reacts with the ε-amine of lysine residues, preserving the cationic nature of the residue; labeled proteins generate extremely sharp peaks in CIEF. A set of four standard proteins generated a linear relationship between migration time and pI. A protein homogenate prepared from a Barrett's esophagus cell line resolved over 100 components in a 40 min separation. Detection limits for Chromeo P503-labeled β-lactoglobulin were 5 amol injected into the capillary. Fluorescent impurities present in the ampholytes generated a large background signal that degraded the detection limit by four orders of magnitude compared with other forms of capillary electrophoresis with this detector.</description><subject>Attomole protein analysis</subject><subject>Calibration</subject><subject>Cell Line</subject><subject>Cells - chemistry</subject><subject>CIEF</subject><subject>Electrophoresis, Capillary - methods</subject><subject>Fluorescent Dyes - chemistry</subject><subject>LIF</subject><subject>Proteins - analysis</subject><subject>Sensitivity and Specificity</subject><subject>Spectrometry, Fluorescence - methods</subject><subject>Time Factors</subject><issn>0173-0835</issn><issn>1522-2683</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2009</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFkEtP4zAURi3ECMpjyxKyYpeO7Ws79hJVbacomhkNINhZTnIDhrQpcSqm_55EqYAdK9_F-Y6sQ8gZo2NGKf-J1TqMOaWaUmH0HhkxyXnMlYZ9MqIsgZhqkIfkKIRn2jNCHJBDZnh_qhERV21bL-sKo3VTt-hXkVu5aht8iLJtNFlMZ9Gbb5-idDGLCmwxb329OiE_SlcFPN29x-RuNr2d_IrTP_PF5CqNcwFKx0mZqRJylTldYsbAyUwx4IzLojS80FyoLKdOcseMTEwhnCoUQo5OCwQm4JhcDt7ub68bDK1d-pBjVbkV1ptglTKCKgPfggBSGwq9cTyAeVOH0GBp141fumZrGbV9UNsHtR9Bu8H5zrzJllh84ruCHWAG4M1XuP1GZ6fp35uv8njY-tDi_4-ta16sSiCR9v733KYivb9-kP_srOMvBr50tXWPjQ_2rtMxoEwapamBd_e1mSE</recordid><startdate>2009</startdate><enddate>2009</enddate><creator>Ramsay, Lauren M</creator><creator>Dickerson, Jane A</creator><creator>Dovichi, Norman J</creator><general>Wiley-VCH Verlag</general><general>WILEY-VCH Verlag</general><general>WILEY‐VCH Verlag</general><scope>FBQ</scope><scope>BSCLL</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7U5</scope><scope>8FD</scope><scope>L7M</scope><scope>7X8</scope></search><sort><creationdate>2009</creationdate><title>Attomole protein analysis by CIEF with LIF detection</title><author>Ramsay, Lauren M ; Dickerson, Jane A ; Dovichi, Norman J</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c4368-7fb6f3c6ba8feb13a5b6132125df92d8246bc0a52a19579d4a6d6e3cea84e3143</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2009</creationdate><topic>Attomole protein analysis</topic><topic>Calibration</topic><topic>Cell Line</topic><topic>Cells - chemistry</topic><topic>CIEF</topic><topic>Electrophoresis, Capillary - methods</topic><topic>Fluorescent Dyes - chemistry</topic><topic>LIF</topic><topic>Proteins - analysis</topic><topic>Sensitivity and Specificity</topic><topic>Spectrometry, Fluorescence - methods</topic><topic>Time Factors</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Ramsay, Lauren M</creatorcontrib><creatorcontrib>Dickerson, Jane A</creatorcontrib><creatorcontrib>Dovichi, Norman J</creatorcontrib><collection>AGRIS</collection><collection>Istex</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Solid State and Superconductivity Abstracts</collection><collection>Technology Research Database</collection><collection>Advanced Technologies Database with Aerospace</collection><collection>MEDLINE - Academic</collection><jtitle>Electrophoresis</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Ramsay, Lauren M</au><au>Dickerson, Jane A</au><au>Dovichi, Norman J</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Attomole protein analysis by CIEF with LIF detection</atitle><jtitle>Electrophoresis</jtitle><addtitle>ELECTROPHORESIS</addtitle><date>2009</date><risdate>2009</risdate><volume>30</volume><issue>2</issue><spage>297</spage><epage>302</epage><pages>297-302</pages><issn>0173-0835</issn><eissn>1522-2683</eissn><abstract>We have coupled CIEF with an LIF detector that is based on a post-column sheath flow cuvette. We employed Chromeo P503 as a fluorogenic reagent to label proteins before analysis. This reagent reacts with the ε-amine of lysine residues, preserving the cationic nature of the residue; labeled proteins generate extremely sharp peaks in CIEF. A set of four standard proteins generated a linear relationship between migration time and pI. A protein homogenate prepared from a Barrett's esophagus cell line resolved over 100 components in a 40 min separation. Detection limits for Chromeo P503-labeled β-lactoglobulin were 5 amol injected into the capillary. Fluorescent impurities present in the ampholytes generated a large background signal that degraded the detection limit by four orders of magnitude compared with other forms of capillary electrophoresis with this detector.</abstract><cop>Weinheim</cop><pub>Wiley-VCH Verlag</pub><pmid>19204946</pmid><doi>10.1002/elps.200800498</doi><tpages>6</tpages></addata></record> |
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| ispartof | Electrophoresis, 2009, Vol.30 (2), p.297-302 |
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| source | MEDLINE; Wiley Online Library Journals Frontfile Complete |
| subjects | Attomole protein analysis Calibration Cell Line Cells - chemistry CIEF Electrophoresis, Capillary - methods Fluorescent Dyes - chemistry LIF Proteins - analysis Sensitivity and Specificity Spectrometry, Fluorescence - methods Time Factors |
| title | Attomole protein analysis by CIEF with LIF detection |
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