Development and validation of novel biomarker assays for osteoarthritis
Osteoarthritis (OA) is the most common chronic joint disease usually diagnosed at relatively advanced stages when there is irreparable damage to the joint(s). Recently, we have identified two novel biomarkers C3f and V65 which appear to be OA-specific and therefore potential markers of early disease...
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| Veröffentlicht in: | PloS one 2017-07, Vol.12 (7), p.e0181334-e0181334 |
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| creator | Ourradi, Khadija Xu, Yunhe de Seny, Dominique Kirwan, John Blom, Ashley Sharif, Mohammed |
| description | Osteoarthritis (OA) is the most common chronic joint disease usually diagnosed at relatively advanced stages when there is irreparable damage to the joint(s). Recently, we have identified two novel biomarkers C3f and V65 which appear to be OA-specific and therefore potential markers of early disease. We report the development of immunoassays for quantitative measure of these two novel biomarkers.
Monoclonal and polyclonal antibodies were generated by immunising mouse and rabbits respectively with peptide-carrier conjugates of C3f and V65. Affinity purified antibodies were used for immunoassays development and assays validated using serum from OA patients and controls.
The ELISAs developed showed spiked recovery of up to 96% for C3f and V65 peptides depending on serum dilutions with a coefficient of variation (CV) |
| doi_str_mv | 10.1371/journal.pone.0181334 |
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Monoclonal and polyclonal antibodies were generated by immunising mouse and rabbits respectively with peptide-carrier conjugates of C3f and V65. Affinity purified antibodies were used for immunoassays development and assays validated using serum from OA patients and controls.
The ELISAs developed showed spiked recovery of up to 96% for C3f and V65 peptides depending on serum dilutions with a coefficient of variation (CV) <10%. The intra- and inter-assay CVs for C3f and V65 were 1.3-10.8% and 4.2-10.3% respectively. Both assays were insensitive for measurements of the peptides in patients and the use of different signal amplification systems did not increase assay sensitivity.
We have developed two immunoassays for measurements of C3f and V65 peptides biomarkers discovered by our earlier proteomic study. These assays could detect the endogenous peptides in serum samples from patients and controls but lacked sensitivity for accurate measurements of the peptides in patients. Our study highlights the difficulties and challenges of validating biomarker from proteomic studies and demonstrates how to overcome some of the technical challenges associated with developing immunoassays for small peptides.</description><identifier>ISSN: 1932-6203</identifier><identifier>EISSN: 1932-6203</identifier><identifier>DOI: 10.1371/journal.pone.0181334</identifier><identifier>PMID: 28715494</identifier><language>eng</language><publisher>United States: Public Library of Science</publisher><subject>Animals ; Antibodies ; Antibody Formation ; Arthritis ; Assaying ; Biocompatibility ; Biological markers ; Biology and Life Sciences ; Biomarkers ; Biomarkers - blood ; Blotting, Western ; Coefficient of variation ; Complement C3b - analysis ; Damage detection ; Diagnosis ; Dilution ; Disease ; Enzyme-Linked Immunosorbent Assay - methods ; Genetic aspects ; Human health sciences ; Humans ; Immunoassay ; Immunoassays ; Mass spectrometry ; Medicine and Health Sciences ; Metabolism ; Mice ; Monoclonal antibodies ; Osteoarthritis ; Osteoarthritis - blood ; Osteoarthritis - diagnosis ; Osteoarthritis - immunology ; Osteoarthritis/blood/diagnosis/immunology ; Patients ; Peptide Fragments - blood ; Peptides ; Physiological aspects ; Polyclonal antibodies ; Proteins ; Rabbits ; Research and Analysis Methods ; Rheumatology ; Rhumatologie ; Sciences de la santé humaine ; Scientific imaging ; Sensitivity ; Sensitivity and Specificity ; Vitronectin - blood</subject><ispartof>PloS one, 2017-07, Vol.12 (7), p.e0181334-e0181334</ispartof><rights>COPYRIGHT 2017 Public Library of Science</rights><rights>2017 Ourradi et al. This is an open access article distributed under the terms of the Creative Commons Attribution License: http://creativecommons.org/licenses/by/4.0/ (the “License”), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Notwithstanding the ProQuest Terms and Conditions, you may use this content in accordance with the terms of the License.</rights><rights>2017 Ourradi et al 2017 Ourradi et al</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c736t-2252589ce2622282c1d2cfb2e99d1a44c56419585b5934368188a464ee52c2383</citedby><cites>FETCH-LOGICAL-c736t-2252589ce2622282c1d2cfb2e99d1a44c56419585b5934368188a464ee52c2383</cites><orcidid>0000-0002-9031-1931</orcidid></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC5513499/pdf/$$EPDF$$P50$$Gpubmedcentral$$Hfree_for_read</linktopdf><linktohtml>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC5513499/$$EHTML$$P50$$Gpubmedcentral$$Hfree_for_read</linktohtml><link.rule.ids>230,314,723,776,780,860,881,2096,2915,23845,27901,27902,53766,53768,79343,79344</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/28715494$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><contributor>D'Auria, Sabato</contributor><creatorcontrib>Ourradi, Khadija</creatorcontrib><creatorcontrib>Xu, Yunhe</creatorcontrib><creatorcontrib>de Seny, Dominique</creatorcontrib><creatorcontrib>Kirwan, John</creatorcontrib><creatorcontrib>Blom, Ashley</creatorcontrib><creatorcontrib>Sharif, Mohammed</creatorcontrib><title>Development and validation of novel biomarker assays for osteoarthritis</title><title>PloS one</title><addtitle>PLoS One</addtitle><description>Osteoarthritis (OA) is the most common chronic joint disease usually diagnosed at relatively advanced stages when there is irreparable damage to the joint(s). Recently, we have identified two novel biomarkers C3f and V65 which appear to be OA-specific and therefore potential markers of early disease. We report the development of immunoassays for quantitative measure of these two novel biomarkers.
Monoclonal and polyclonal antibodies were generated by immunising mouse and rabbits respectively with peptide-carrier conjugates of C3f and V65. Affinity purified antibodies were used for immunoassays development and assays validated using serum from OA patients and controls.
The ELISAs developed showed spiked recovery of up to 96% for C3f and V65 peptides depending on serum dilutions with a coefficient of variation (CV) <10%. The intra- and inter-assay CVs for C3f and V65 were 1.3-10.8% and 4.2-10.3% respectively. Both assays were insensitive for measurements of the peptides in patients and the use of different signal amplification systems did not increase assay sensitivity.
We have developed two immunoassays for measurements of C3f and V65 peptides biomarkers discovered by our earlier proteomic study. These assays could detect the endogenous peptides in serum samples from patients and controls but lacked sensitivity for accurate measurements of the peptides in patients. Our study highlights the difficulties and challenges of validating biomarker from proteomic studies and demonstrates how to overcome some of the technical challenges associated with developing immunoassays for small peptides.</description><subject>Animals</subject><subject>Antibodies</subject><subject>Antibody Formation</subject><subject>Arthritis</subject><subject>Assaying</subject><subject>Biocompatibility</subject><subject>Biological markers</subject><subject>Biology and Life Sciences</subject><subject>Biomarkers</subject><subject>Biomarkers - blood</subject><subject>Blotting, Western</subject><subject>Coefficient of variation</subject><subject>Complement C3b - analysis</subject><subject>Damage detection</subject><subject>Diagnosis</subject><subject>Dilution</subject><subject>Disease</subject><subject>Enzyme-Linked Immunosorbent Assay - methods</subject><subject>Genetic aspects</subject><subject>Human health sciences</subject><subject>Humans</subject><subject>Immunoassay</subject><subject>Immunoassays</subject><subject>Mass spectrometry</subject><subject>Medicine and Health Sciences</subject><subject>Metabolism</subject><subject>Mice</subject><subject>Monoclonal antibodies</subject><subject>Osteoarthritis</subject><subject>Osteoarthritis - blood</subject><subject>Osteoarthritis - diagnosis</subject><subject>Osteoarthritis - immunology</subject><subject>Osteoarthritis/blood/diagnosis/immunology</subject><subject>Patients</subject><subject>Peptide Fragments - blood</subject><subject>Peptides</subject><subject>Physiological aspects</subject><subject>Polyclonal antibodies</subject><subject>Proteins</subject><subject>Rabbits</subject><subject>Research and Analysis Methods</subject><subject>Rheumatology</subject><subject>Rhumatologie</subject><subject>Sciences de la santé humaine</subject><subject>Scientific imaging</subject><subject>Sensitivity</subject><subject>Sensitivity and Specificity</subject><subject>Vitronectin - blood</subject><issn>1932-6203</issn><issn>1932-6203</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2017</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><sourceid>BENPR</sourceid><sourceid>DOA</sourceid><recordid>eNqNU11v0zAUjRCIjcE_QBAJCcFDiz8T-wVpGjAqTZrE16vlODetixt3dlKxf4_TZlOD9oDyEMs-59xzj32z7CVGc0xL_GHt-9BqN9_6FuYIC0wpe5SdYknJrCCIPj5an2TPYlwjxKkoiqfZCREl5kyy0-zyE-zA-e0G2i7XbZ3vtLO17qxvc9_krU-neWX9RoffEHIdo76NeeND7mMHXoduFWxn4_PsSaNdhBfj_yz7-eXzj4uvs6vry8XF-dXMlLToZoRwwoU0QApCiCAG18Q0FQEpa6wZM7xgWHLBKy4po4XAQmhWMABODKGCnmWvD7pb56MaM4gKS4k5x4ihhFgcELXXa7UNNlm_VV5btd_wYamSa2scKI2ExLLiWFeCcYpkASDrUpayqSva4KT1cazWVxuoTQopaDcRnZ60dqWWfqeSF8qkTAL0IOAsLCEVr6zakT1xv-5dcmNUBYqQQqihQ0oS691YNvibHmKnNjYacE634PuhW1JSlCIc8njzD_ThTEbUUqe2bdv45NYMouqcI1wWiLDB7PwBVPpq2FiTnllj0_6E8H5CSJgO_nRL3ceoFt-__T_2-tcU-_YIuwLtulX0rh9eZZwC2QFogo8xQHN_NRipYUru0lDDlKhxShLt1fG13pPuxoL-BbnxCjU</recordid><startdate>20170717</startdate><enddate>20170717</enddate><creator>Ourradi, Khadija</creator><creator>Xu, Yunhe</creator><creator>de Seny, Dominique</creator><creator>Kirwan, John</creator><creator>Blom, Ashley</creator><creator>Sharif, Mohammed</creator><general>Public Library of Science</general><general>Public Library of Science (PLoS)</general><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>IOV</scope><scope>ISR</scope><scope>3V.</scope><scope>7QG</scope><scope>7QL</scope><scope>7QO</scope><scope>7RV</scope><scope>7SN</scope><scope>7SS</scope><scope>7T5</scope><scope>7TG</scope><scope>7TM</scope><scope>7U9</scope><scope>7X2</scope><scope>7X7</scope><scope>7XB</scope><scope>88E</scope><scope>8AO</scope><scope>8C1</scope><scope>8FD</scope><scope>8FE</scope><scope>8FG</scope><scope>8FH</scope><scope>8FI</scope><scope>8FJ</scope><scope>8FK</scope><scope>ABJCF</scope><scope>ABUWG</scope><scope>AEUYN</scope><scope>AFKRA</scope><scope>ARAPS</scope><scope>ATCPS</scope><scope>AZQEC</scope><scope>BBNVY</scope><scope>BENPR</scope><scope>BGLVJ</scope><scope>BHPHI</scope><scope>C1K</scope><scope>CCPQU</scope><scope>D1I</scope><scope>DWQXO</scope><scope>FR3</scope><scope>FYUFA</scope><scope>GHDGH</scope><scope>GNUQQ</scope><scope>H94</scope><scope>HCIFZ</scope><scope>K9.</scope><scope>KB.</scope><scope>KB0</scope><scope>KL.</scope><scope>L6V</scope><scope>LK8</scope><scope>M0K</scope><scope>M0S</scope><scope>M1P</scope><scope>M7N</scope><scope>M7P</scope><scope>M7S</scope><scope>NAPCQ</scope><scope>P5Z</scope><scope>P62</scope><scope>P64</scope><scope>PATMY</scope><scope>PDBOC</scope><scope>PIMPY</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>PRINS</scope><scope>PTHSS</scope><scope>PYCSY</scope><scope>RC3</scope><scope>7X8</scope><scope>Q33</scope><scope>5PM</scope><scope>DOA</scope><orcidid>https://orcid.org/0000-0002-9031-1931</orcidid></search><sort><creationdate>20170717</creationdate><title>Development and validation of novel biomarker assays for osteoarthritis</title><author>Ourradi, Khadija ; Xu, Yunhe ; de Seny, Dominique ; Kirwan, John ; Blom, Ashley ; Sharif, Mohammed</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c736t-2252589ce2622282c1d2cfb2e99d1a44c56419585b5934368188a464ee52c2383</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2017</creationdate><topic>Animals</topic><topic>Antibodies</topic><topic>Antibody Formation</topic><topic>Arthritis</topic><topic>Assaying</topic><topic>Biocompatibility</topic><topic>Biological markers</topic><topic>Biology and Life Sciences</topic><topic>Biomarkers</topic><topic>Biomarkers - 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Academic</collection><collection>Université de Liège - Open Repository and Bibliography (ORBI)</collection><collection>PubMed Central (Full Participant titles)</collection><collection>DOAJ Directory of Open Access Journals</collection><jtitle>PloS one</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Ourradi, Khadija</au><au>Xu, Yunhe</au><au>de Seny, Dominique</au><au>Kirwan, John</au><au>Blom, Ashley</au><au>Sharif, Mohammed</au><au>D'Auria, Sabato</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Development and validation of novel biomarker assays for osteoarthritis</atitle><jtitle>PloS one</jtitle><addtitle>PLoS One</addtitle><date>2017-07-17</date><risdate>2017</risdate><volume>12</volume><issue>7</issue><spage>e0181334</spage><epage>e0181334</epage><pages>e0181334-e0181334</pages><issn>1932-6203</issn><eissn>1932-6203</eissn><abstract>Osteoarthritis (OA) is the most common chronic joint disease usually diagnosed at relatively advanced stages when there is irreparable damage to the joint(s). Recently, we have identified two novel biomarkers C3f and V65 which appear to be OA-specific and therefore potential markers of early disease. We report the development of immunoassays for quantitative measure of these two novel biomarkers.
Monoclonal and polyclonal antibodies were generated by immunising mouse and rabbits respectively with peptide-carrier conjugates of C3f and V65. Affinity purified antibodies were used for immunoassays development and assays validated using serum from OA patients and controls.
The ELISAs developed showed spiked recovery of up to 96% for C3f and V65 peptides depending on serum dilutions with a coefficient of variation (CV) <10%. The intra- and inter-assay CVs for C3f and V65 were 1.3-10.8% and 4.2-10.3% respectively. Both assays were insensitive for measurements of the peptides in patients and the use of different signal amplification systems did not increase assay sensitivity.
We have developed two immunoassays for measurements of C3f and V65 peptides biomarkers discovered by our earlier proteomic study. These assays could detect the endogenous peptides in serum samples from patients and controls but lacked sensitivity for accurate measurements of the peptides in patients. Our study highlights the difficulties and challenges of validating biomarker from proteomic studies and demonstrates how to overcome some of the technical challenges associated with developing immunoassays for small peptides.</abstract><cop>United States</cop><pub>Public Library of Science</pub><pmid>28715494</pmid><doi>10.1371/journal.pone.0181334</doi><tpages>e0181334</tpages><orcidid>https://orcid.org/0000-0002-9031-1931</orcidid><oa>free_for_read</oa></addata></record> |
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| source | Public Library of Science (PLoS) Journals Open Access; MEDLINE; DOAJ Directory of Open Access Journals; EZB-FREE-00999 freely available EZB journals; PubMed Central; Free Full-Text Journals in Chemistry |
| subjects | Animals Antibodies Antibody Formation Arthritis Assaying Biocompatibility Biological markers Biology and Life Sciences Biomarkers Biomarkers - blood Blotting, Western Coefficient of variation Complement C3b - analysis Damage detection Diagnosis Dilution Disease Enzyme-Linked Immunosorbent Assay - methods Genetic aspects Human health sciences Humans Immunoassay Immunoassays Mass spectrometry Medicine and Health Sciences Metabolism Mice Monoclonal antibodies Osteoarthritis Osteoarthritis - blood Osteoarthritis - diagnosis Osteoarthritis - immunology Osteoarthritis/blood/diagnosis/immunology Patients Peptide Fragments - blood Peptides Physiological aspects Polyclonal antibodies Proteins Rabbits Research and Analysis Methods Rheumatology Rhumatologie Sciences de la santé humaine Scientific imaging Sensitivity Sensitivity and Specificity Vitronectin - blood |
| title | Development and validation of novel biomarker assays for osteoarthritis |
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