An approach to remove albumin for the proteomic analysis of low abundance biomarkers in human serum

Proteomic technologies are being used to discover and identify disease‐associated biomarkers. The application of these technologies in the search for potential diagnostic/prognostic biomarkers in the serum of patients has been limited by the presence of highly abundant albumin and immunoglobulins th...

Ausführliche Beschreibung

Gespeichert in:
Bibliographische Detailangaben
Veröffentlicht in:Proteomics (Weinheim) 2003-10, Vol.3 (10), p.1980-1987
Hauptverfasser: Ahmed, Nuzhat, Barker, Gillian, Oliva, Karen, Garfin, David, Talmadge, Kenneth, Georgiou, Harry, Quinn, Michael, Rice, Greg
Format: Artikel
Sprache:eng
Schlagworte:
Online-Zugang:Volltext
Tags: Tag hinzufügen
Keine Tags, Fügen Sie den ersten Tag hinzu!
container_end_page 1987
container_issue 10
container_start_page 1980
container_title Proteomics (Weinheim)
container_volume 3
creator Ahmed, Nuzhat
Barker, Gillian
Oliva, Karen
Garfin, David
Talmadge, Kenneth
Georgiou, Harry
Quinn, Michael
Rice, Greg
description Proteomic technologies are being used to discover and identify disease‐associated biomarkers. The application of these technologies in the search for potential diagnostic/prognostic biomarkers in the serum of patients has been limited by the presence of highly abundant albumin and immunoglobulins that constitute approximately 60–97% of the total serum proteins. The purpose of the study was to evaluate whether treatment of human serum with Affi‐Gel Blue alone or in combination with Protein A (Aurum serum protein mini kit, Bio‐Rad) before two‐dimensional gel electrophoresis (2‐DE) analysis removed high abundance proteins to allow the visualization of low abundant proteins. Serum samples were treated with either Affi‐Gel Blue or Aurum kit and then subjected to 2‐DE using 11 cm, pH 4–7 isoelectric focussing strips for the first dimension and 10% sodium dodecyl sulfate‐polyacrylamide gel electrophoresis for second dimension. Protein spots were visualized using a fluorescent protein dye (SYPRO Ruby, Bio‐Rad). Comparison between treatment methods showed significant removal of albumin by both Affi‐Gel Blue and Aurum kit and considerable differences in the protein profile of the gels after each treatment. Direct comparison between treatments revealed twenty‐eight protein spots unique to Affi‐Gel Blue while only two spots were unique after Aurum kit treatment. Unique spots in Affi‐Gel Blue and Aurum kit treated serum were not visualized in untreated serum. Sixteen hours of Affi‐Gel Blue treatment resulted in enhanced visualization of fifty‐three protein spots by two‐fold, thirty‐one by five‐fold, twelve by ten‐fold and six by twenty‐fold. In parallel after Aurum kit treatment two‐, five‐, ten‐ and twenty‐fold enhancements of thirty, thirteen, eight and five protein spots, respectively, were observed. The pattern of increased visualization of protein spots with both treatment methods was similar. In conclusion, treatment of serum samples with Affi‐Gel Blue or Aurum kit before 2‐DE analysis can be used to remove high abundance proteins in order to increase the detection sensitivity of proteins present in low abundance.
doi_str_mv 10.1002/pmic.200300465
format Article
fullrecord <record><control><sourceid>proquest_cross</sourceid><recordid>TN_cdi_proquest_miscellaneous_71421976</recordid><sourceformat>XML</sourceformat><sourcesystem>PC</sourcesystem><sourcerecordid>71421976</sourcerecordid><originalsourceid>FETCH-LOGICAL-c3795-39df30cde9caad21c8435feef1e6eaa1e12eb4d0b6415abcaa8448bf884ff4173</originalsourceid><addsrcrecordid>eNqFkE1P3DAQhi1ExVe5ckQ-9ZbFEzt2ckSrsiCWfkit2pvlOGNtaBJv7QTYf1-jXS29cZo5PO87o4eQC2AzYCy_WvetneWMccaELA7ICUgosqqUcLjfC35MTmN8ZAxUWakjcgxC5kUp2Qmx1wM163Xwxq7o6GnA3j8hNV099e1AnQ90XCFNwIg-3aJmMN0mtpF6Rzv_TE09DY0ZLNK69b0JfzBEmpKrqTcDjRim_iP54EwX8Xw3z8jPm88_5rfZ8uvibn69zCxXVZHxqnGc2QYra0yTgy0FLxyiA5RoDCDkWIuG1VJAYeoElUKUtStL4ZwAxc_Ip21v-vbvhHHUfRstdp0Z0E9RKxA5VEomcLYFbfAxBnR6Hdr0-0YD069a9atWvdeaApe75qnusXnDdx4TUG2B57bDzTt1-tvD3fz_8mybbeOIL_tsUqml4qrQv74s9FLJ29_330Hn_B9Vp5W_</addsrcrecordid><sourcetype>Aggregation Database</sourcetype><iscdi>true</iscdi><recordtype>article</recordtype><pqid>71421976</pqid></control><display><type>article</type><title>An approach to remove albumin for the proteomic analysis of low abundance biomarkers in human serum</title><source>MEDLINE</source><source>Access via Wiley Online Library</source><creator>Ahmed, Nuzhat ; Barker, Gillian ; Oliva, Karen ; Garfin, David ; Talmadge, Kenneth ; Georgiou, Harry ; Quinn, Michael ; Rice, Greg</creator><creatorcontrib>Ahmed, Nuzhat ; Barker, Gillian ; Oliva, Karen ; Garfin, David ; Talmadge, Kenneth ; Georgiou, Harry ; Quinn, Michael ; Rice, Greg</creatorcontrib><description>Proteomic technologies are being used to discover and identify disease‐associated biomarkers. The application of these technologies in the search for potential diagnostic/prognostic biomarkers in the serum of patients has been limited by the presence of highly abundant albumin and immunoglobulins that constitute approximately 60–97% of the total serum proteins. The purpose of the study was to evaluate whether treatment of human serum with Affi‐Gel Blue alone or in combination with Protein A (Aurum serum protein mini kit, Bio‐Rad) before two‐dimensional gel electrophoresis (2‐DE) analysis removed high abundance proteins to allow the visualization of low abundant proteins. Serum samples were treated with either Affi‐Gel Blue or Aurum kit and then subjected to 2‐DE using 11 cm, pH 4–7 isoelectric focussing strips for the first dimension and 10% sodium dodecyl sulfate‐polyacrylamide gel electrophoresis for second dimension. Protein spots were visualized using a fluorescent protein dye (SYPRO Ruby, Bio‐Rad). Comparison between treatment methods showed significant removal of albumin by both Affi‐Gel Blue and Aurum kit and considerable differences in the protein profile of the gels after each treatment. Direct comparison between treatments revealed twenty‐eight protein spots unique to Affi‐Gel Blue while only two spots were unique after Aurum kit treatment. Unique spots in Affi‐Gel Blue and Aurum kit treated serum were not visualized in untreated serum. Sixteen hours of Affi‐Gel Blue treatment resulted in enhanced visualization of fifty‐three protein spots by two‐fold, thirty‐one by five‐fold, twelve by ten‐fold and six by twenty‐fold. In parallel after Aurum kit treatment two‐, five‐, ten‐ and twenty‐fold enhancements of thirty, thirteen, eight and five protein spots, respectively, were observed. The pattern of increased visualization of protein spots with both treatment methods was similar. In conclusion, treatment of serum samples with Affi‐Gel Blue or Aurum kit before 2‐DE analysis can be used to remove high abundance proteins in order to increase the detection sensitivity of proteins present in low abundance.</description><identifier>ISSN: 1615-9853</identifier><identifier>EISSN: 1615-9861</identifier><identifier>DOI: 10.1002/pmic.200300465</identifier><identifier>PMID: 14625860</identifier><language>eng</language><publisher>Weinheim: WILEY-VCH Verlag</publisher><subject>Affi-Gel Blue ; Albumin ; Aurum kit ; Biomarkers ; Biomarkers - blood ; Blood Protein Electrophoresis ; Blood Proteins - analysis ; Blood Proteins - isolation &amp; purification ; Chromatography, Affinity ; Electrophoresis, Gel, Two-Dimensional ; Humans ; Immunoglobulin ; Isoelectric Focusing ; Proteome - analysis ; Proteomics - methods ; Serum Albumin - isolation &amp; purification ; Time Factors ; Triazines - chemistry ; Two-dimensional gel electrophoresis</subject><ispartof>Proteomics (Weinheim), 2003-10, Vol.3 (10), p.1980-1987</ispartof><rights>Copyright © 2003 WILEY‐VCH Verlag GmbH &amp; Co. KGaA, Weinheim</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c3795-39df30cde9caad21c8435feef1e6eaa1e12eb4d0b6415abcaa8448bf884ff4173</citedby><cites>FETCH-LOGICAL-c3795-39df30cde9caad21c8435feef1e6eaa1e12eb4d0b6415abcaa8448bf884ff4173</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://onlinelibrary.wiley.com/doi/pdf/10.1002%2Fpmic.200300465$$EPDF$$P50$$Gwiley$$H</linktopdf><linktohtml>$$Uhttps://onlinelibrary.wiley.com/doi/full/10.1002%2Fpmic.200300465$$EHTML$$P50$$Gwiley$$H</linktohtml><link.rule.ids>314,780,784,1417,27924,27925,45574,45575</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/14625860$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Ahmed, Nuzhat</creatorcontrib><creatorcontrib>Barker, Gillian</creatorcontrib><creatorcontrib>Oliva, Karen</creatorcontrib><creatorcontrib>Garfin, David</creatorcontrib><creatorcontrib>Talmadge, Kenneth</creatorcontrib><creatorcontrib>Georgiou, Harry</creatorcontrib><creatorcontrib>Quinn, Michael</creatorcontrib><creatorcontrib>Rice, Greg</creatorcontrib><title>An approach to remove albumin for the proteomic analysis of low abundance biomarkers in human serum</title><title>Proteomics (Weinheim)</title><addtitle>Proteomics</addtitle><description>Proteomic technologies are being used to discover and identify disease‐associated biomarkers. The application of these technologies in the search for potential diagnostic/prognostic biomarkers in the serum of patients has been limited by the presence of highly abundant albumin and immunoglobulins that constitute approximately 60–97% of the total serum proteins. The purpose of the study was to evaluate whether treatment of human serum with Affi‐Gel Blue alone or in combination with Protein A (Aurum serum protein mini kit, Bio‐Rad) before two‐dimensional gel electrophoresis (2‐DE) analysis removed high abundance proteins to allow the visualization of low abundant proteins. Serum samples were treated with either Affi‐Gel Blue or Aurum kit and then subjected to 2‐DE using 11 cm, pH 4–7 isoelectric focussing strips for the first dimension and 10% sodium dodecyl sulfate‐polyacrylamide gel electrophoresis for second dimension. Protein spots were visualized using a fluorescent protein dye (SYPRO Ruby, Bio‐Rad). Comparison between treatment methods showed significant removal of albumin by both Affi‐Gel Blue and Aurum kit and considerable differences in the protein profile of the gels after each treatment. Direct comparison between treatments revealed twenty‐eight protein spots unique to Affi‐Gel Blue while only two spots were unique after Aurum kit treatment. Unique spots in Affi‐Gel Blue and Aurum kit treated serum were not visualized in untreated serum. Sixteen hours of Affi‐Gel Blue treatment resulted in enhanced visualization of fifty‐three protein spots by two‐fold, thirty‐one by five‐fold, twelve by ten‐fold and six by twenty‐fold. In parallel after Aurum kit treatment two‐, five‐, ten‐ and twenty‐fold enhancements of thirty, thirteen, eight and five protein spots, respectively, were observed. The pattern of increased visualization of protein spots with both treatment methods was similar. In conclusion, treatment of serum samples with Affi‐Gel Blue or Aurum kit before 2‐DE analysis can be used to remove high abundance proteins in order to increase the detection sensitivity of proteins present in low abundance.</description><subject>Affi-Gel Blue</subject><subject>Albumin</subject><subject>Aurum kit</subject><subject>Biomarkers</subject><subject>Biomarkers - blood</subject><subject>Blood Protein Electrophoresis</subject><subject>Blood Proteins - analysis</subject><subject>Blood Proteins - isolation &amp; purification</subject><subject>Chromatography, Affinity</subject><subject>Electrophoresis, Gel, Two-Dimensional</subject><subject>Humans</subject><subject>Immunoglobulin</subject><subject>Isoelectric Focusing</subject><subject>Proteome - analysis</subject><subject>Proteomics - methods</subject><subject>Serum Albumin - isolation &amp; purification</subject><subject>Time Factors</subject><subject>Triazines - chemistry</subject><subject>Two-dimensional gel electrophoresis</subject><issn>1615-9853</issn><issn>1615-9861</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2003</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFkE1P3DAQhi1ExVe5ckQ-9ZbFEzt2ckSrsiCWfkit2pvlOGNtaBJv7QTYf1-jXS29cZo5PO87o4eQC2AzYCy_WvetneWMccaELA7ICUgosqqUcLjfC35MTmN8ZAxUWakjcgxC5kUp2Qmx1wM163Xwxq7o6GnA3j8hNV099e1AnQ90XCFNwIg-3aJmMN0mtpF6Rzv_TE09DY0ZLNK69b0JfzBEmpKrqTcDjRim_iP54EwX8Xw3z8jPm88_5rfZ8uvibn69zCxXVZHxqnGc2QYra0yTgy0FLxyiA5RoDCDkWIuG1VJAYeoElUKUtStL4ZwAxc_Ip21v-vbvhHHUfRstdp0Z0E9RKxA5VEomcLYFbfAxBnR6Hdr0-0YD069a9atWvdeaApe75qnusXnDdx4TUG2B57bDzTt1-tvD3fz_8mybbeOIL_tsUqml4qrQv74s9FLJ29_330Hn_B9Vp5W_</recordid><startdate>200310</startdate><enddate>200310</enddate><creator>Ahmed, Nuzhat</creator><creator>Barker, Gillian</creator><creator>Oliva, Karen</creator><creator>Garfin, David</creator><creator>Talmadge, Kenneth</creator><creator>Georgiou, Harry</creator><creator>Quinn, Michael</creator><creator>Rice, Greg</creator><general>WILEY-VCH Verlag</general><general>WILEY‐VCH Verlag</general><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>7X8</scope></search><sort><creationdate>200310</creationdate><title>An approach to remove albumin for the proteomic analysis of low abundance biomarkers in human serum</title><author>Ahmed, Nuzhat ; Barker, Gillian ; Oliva, Karen ; Garfin, David ; Talmadge, Kenneth ; Georgiou, Harry ; Quinn, Michael ; Rice, Greg</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c3795-39df30cde9caad21c8435feef1e6eaa1e12eb4d0b6415abcaa8448bf884ff4173</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2003</creationdate><topic>Affi-Gel Blue</topic><topic>Albumin</topic><topic>Aurum kit</topic><topic>Biomarkers</topic><topic>Biomarkers - blood</topic><topic>Blood Protein Electrophoresis</topic><topic>Blood Proteins - analysis</topic><topic>Blood Proteins - isolation &amp; purification</topic><topic>Chromatography, Affinity</topic><topic>Electrophoresis, Gel, Two-Dimensional</topic><topic>Humans</topic><topic>Immunoglobulin</topic><topic>Isoelectric Focusing</topic><topic>Proteome - analysis</topic><topic>Proteomics - methods</topic><topic>Serum Albumin - isolation &amp; purification</topic><topic>Time Factors</topic><topic>Triazines - chemistry</topic><topic>Two-dimensional gel electrophoresis</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Ahmed, Nuzhat</creatorcontrib><creatorcontrib>Barker, Gillian</creatorcontrib><creatorcontrib>Oliva, Karen</creatorcontrib><creatorcontrib>Garfin, David</creatorcontrib><creatorcontrib>Talmadge, Kenneth</creatorcontrib><creatorcontrib>Georgiou, Harry</creatorcontrib><creatorcontrib>Quinn, Michael</creatorcontrib><creatorcontrib>Rice, Greg</creatorcontrib><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>MEDLINE - Academic</collection><jtitle>Proteomics (Weinheim)</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Ahmed, Nuzhat</au><au>Barker, Gillian</au><au>Oliva, Karen</au><au>Garfin, David</au><au>Talmadge, Kenneth</au><au>Georgiou, Harry</au><au>Quinn, Michael</au><au>Rice, Greg</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>An approach to remove albumin for the proteomic analysis of low abundance biomarkers in human serum</atitle><jtitle>Proteomics (Weinheim)</jtitle><addtitle>Proteomics</addtitle><date>2003-10</date><risdate>2003</risdate><volume>3</volume><issue>10</issue><spage>1980</spage><epage>1987</epage><pages>1980-1987</pages><issn>1615-9853</issn><eissn>1615-9861</eissn><abstract>Proteomic technologies are being used to discover and identify disease‐associated biomarkers. The application of these technologies in the search for potential diagnostic/prognostic biomarkers in the serum of patients has been limited by the presence of highly abundant albumin and immunoglobulins that constitute approximately 60–97% of the total serum proteins. The purpose of the study was to evaluate whether treatment of human serum with Affi‐Gel Blue alone or in combination with Protein A (Aurum serum protein mini kit, Bio‐Rad) before two‐dimensional gel electrophoresis (2‐DE) analysis removed high abundance proteins to allow the visualization of low abundant proteins. Serum samples were treated with either Affi‐Gel Blue or Aurum kit and then subjected to 2‐DE using 11 cm, pH 4–7 isoelectric focussing strips for the first dimension and 10% sodium dodecyl sulfate‐polyacrylamide gel electrophoresis for second dimension. Protein spots were visualized using a fluorescent protein dye (SYPRO Ruby, Bio‐Rad). Comparison between treatment methods showed significant removal of albumin by both Affi‐Gel Blue and Aurum kit and considerable differences in the protein profile of the gels after each treatment. Direct comparison between treatments revealed twenty‐eight protein spots unique to Affi‐Gel Blue while only two spots were unique after Aurum kit treatment. Unique spots in Affi‐Gel Blue and Aurum kit treated serum were not visualized in untreated serum. Sixteen hours of Affi‐Gel Blue treatment resulted in enhanced visualization of fifty‐three protein spots by two‐fold, thirty‐one by five‐fold, twelve by ten‐fold and six by twenty‐fold. In parallel after Aurum kit treatment two‐, five‐, ten‐ and twenty‐fold enhancements of thirty, thirteen, eight and five protein spots, respectively, were observed. The pattern of increased visualization of protein spots with both treatment methods was similar. In conclusion, treatment of serum samples with Affi‐Gel Blue or Aurum kit before 2‐DE analysis can be used to remove high abundance proteins in order to increase the detection sensitivity of proteins present in low abundance.</abstract><cop>Weinheim</cop><pub>WILEY-VCH Verlag</pub><pmid>14625860</pmid><doi>10.1002/pmic.200300465</doi><tpages>8</tpages></addata></record>
fulltext fulltext
identifier ISSN: 1615-9853
ispartof Proteomics (Weinheim), 2003-10, Vol.3 (10), p.1980-1987
issn 1615-9853
1615-9861
language eng
recordid cdi_proquest_miscellaneous_71421976
source MEDLINE; Access via Wiley Online Library
subjects Affi-Gel Blue
Albumin
Aurum kit
Biomarkers
Biomarkers - blood
Blood Protein Electrophoresis
Blood Proteins - analysis
Blood Proteins - isolation & purification
Chromatography, Affinity
Electrophoresis, Gel, Two-Dimensional
Humans
Immunoglobulin
Isoelectric Focusing
Proteome - analysis
Proteomics - methods
Serum Albumin - isolation & purification
Time Factors
Triazines - chemistry
Two-dimensional gel electrophoresis
title An approach to remove albumin for the proteomic analysis of low abundance biomarkers in human serum
url https://sfx.bib-bvb.de/sfx_tum?ctx_ver=Z39.88-2004&ctx_enc=info:ofi/enc:UTF-8&ctx_tim=2024-12-25T21%3A53%3A32IST&url_ver=Z39.88-2004&url_ctx_fmt=infofi/fmt:kev:mtx:ctx&rfr_id=info:sid/primo.exlibrisgroup.com:primo3-Article-proquest_cross&rft_val_fmt=info:ofi/fmt:kev:mtx:journal&rft.genre=article&rft.atitle=An%20approach%20to%20remove%20albumin%20for%20the%20proteomic%20analysis%20of%20low%20abundance%20biomarkers%20in%20human%20serum&rft.jtitle=Proteomics%20(Weinheim)&rft.au=Ahmed,%20Nuzhat&rft.date=2003-10&rft.volume=3&rft.issue=10&rft.spage=1980&rft.epage=1987&rft.pages=1980-1987&rft.issn=1615-9853&rft.eissn=1615-9861&rft_id=info:doi/10.1002/pmic.200300465&rft_dat=%3Cproquest_cross%3E71421976%3C/proquest_cross%3E%3Curl%3E%3C/url%3E&disable_directlink=true&sfx.directlink=off&sfx.report_link=0&rft_id=info:oai/&rft_pqid=71421976&rft_id=info:pmid/14625860&rfr_iscdi=true