Development of glutamic acid decarboxylase 65 (GAD65) autoantibody assay using biotin–GAD65 fusion protein
We evaluated a biotin–glutamic acid decarboxylase 65 (GAD65)-based enzyme-linked immunosorbent assay (B-ELISA) to detect GAD65 autoantibodies (GAD65Ab) in 78 sera from individuals with newly diagnosed type 1 diabetes. The GAD65Ab index of patients with type 1 diabetes (mean value of GAD65Ab index of...
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| Veröffentlicht in: | Journal of biotechnology 2004-07, Vol.111 (1), p.97-104 |
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| creator | Luo, Dong Rogers, Chad N Steed, Jordan T Gilliam, Lisa K Hampe, Christiane S Lernmark, Åke |
| description | We evaluated a biotin–glutamic acid decarboxylase 65 (GAD65)-based enzyme-linked immunosorbent assay (B-ELISA) to detect GAD65 autoantibodies (GAD65Ab) in 78 sera from individuals with newly diagnosed type 1 diabetes. The GAD65Ab index of patients with type 1 diabetes (mean value of GAD65Ab index of 1.891) was significantly higher than those in 50 sera from healthy control group (mean value of 0.068). The intra- and inter-assay coefficients of variation (CV) were calculated to be 1.042 and 10.703%, respectively. The specificity of the B-GAD65 ELISA was comparable to the standard radioimmunoassay (RIA) which is routinely used in the laboratory. We describe the optimal conditions of the binding kinetics from each assay-step for the detection of GAD65Ab using the WHO standard serum 97/550 as a model autoantibody serum. We concluded that incubation times of 15, 90, and 90
min for step 1 (pre-incubation of Biotin14–GAD65 with serum), step 2 (binding the Ab/Ag complex to NeutrAvidin plate), and step 3 (incubation with HRPO-anti-human IgG), respectively, along with human serum dilutions of 1:50, would provide an optimal assay signal within a relatively short timeframe. |
| doi_str_mv | 10.1016/j.jbiotec.2004.03.009 |
| format | Article |
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min for step 1 (pre-incubation of Biotin14–GAD65 with serum), step 2 (binding the Ab/Ag complex to NeutrAvidin plate), and step 3 (incubation with HRPO-anti-human IgG), respectively, along with human serum dilutions of 1:50, would provide an optimal assay signal within a relatively short timeframe.</description><identifier>ISSN: 0168-1656</identifier><identifier>EISSN: 1873-4863</identifier><identifier>DOI: 10.1016/j.jbiotec.2004.03.009</identifier><identifier>PMID: 15196774</identifier><identifier>CODEN: JBITD4</identifier><language>eng</language><publisher>Lausanne: Elsevier B.V</publisher><subject>Antibody binding assays ; Autoantigens - blood ; Autoantigens - immunology ; Biological and medical sciences ; Biomarkers - blood ; Biotechnology ; Biotin - immunology ; Biotin–GAD65 ; Blood Chemical Analysis - methods ; Diabetes Mellitus, Type 1 - blood ; Diabetes Mellitus, Type 1 - diagnosis ; Diabetes Mellitus, Type 1 - immunology ; Enzyme-Linked Immunosorbent Assay - methods ; Fundamental and applied biological sciences. Psychology ; Glutamate Decarboxylase - blood ; Glutamate Decarboxylase - immunology ; Humans ; Recombinant Fusion Proteins ; Reproducibility of Results ; Sensitivity and Specificity ; Type 1 diabetes</subject><ispartof>Journal of biotechnology, 2004-07, Vol.111 (1), p.97-104</ispartof><rights>2004 Elsevier B.V.</rights><rights>2004 INIST-CNRS</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c459t-7376ab80a73f5ef6a9653fb07872c83da02e98e5cc74d6b6ee3247e227632bf03</citedby><cites>FETCH-LOGICAL-c459t-7376ab80a73f5ef6a9653fb07872c83da02e98e5cc74d6b6ee3247e227632bf03</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktohtml>$$Uhttps://dx.doi.org/10.1016/j.jbiotec.2004.03.009$$EHTML$$P50$$Gelsevier$$H</linktohtml><link.rule.ids>314,780,784,3550,27924,27925,45995</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=15867288$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/15196774$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Luo, Dong</creatorcontrib><creatorcontrib>Rogers, Chad N</creatorcontrib><creatorcontrib>Steed, Jordan T</creatorcontrib><creatorcontrib>Gilliam, Lisa K</creatorcontrib><creatorcontrib>Hampe, Christiane S</creatorcontrib><creatorcontrib>Lernmark, Åke</creatorcontrib><title>Development of glutamic acid decarboxylase 65 (GAD65) autoantibody assay using biotin–GAD65 fusion protein</title><title>Journal of biotechnology</title><addtitle>J Biotechnol</addtitle><description>We evaluated a biotin–glutamic acid decarboxylase 65 (GAD65)-based enzyme-linked immunosorbent assay (B-ELISA) to detect GAD65 autoantibodies (GAD65Ab) in 78 sera from individuals with newly diagnosed type 1 diabetes. The GAD65Ab index of patients with type 1 diabetes (mean value of GAD65Ab index of 1.891) was significantly higher than those in 50 sera from healthy control group (mean value of 0.068). The intra- and inter-assay coefficients of variation (CV) were calculated to be 1.042 and 10.703%, respectively. The specificity of the B-GAD65 ELISA was comparable to the standard radioimmunoassay (RIA) which is routinely used in the laboratory. We describe the optimal conditions of the binding kinetics from each assay-step for the detection of GAD65Ab using the WHO standard serum 97/550 as a model autoantibody serum. We concluded that incubation times of 15, 90, and 90
min for step 1 (pre-incubation of Biotin14–GAD65 with serum), step 2 (binding the Ab/Ag complex to NeutrAvidin plate), and step 3 (incubation with HRPO-anti-human IgG), respectively, along with human serum dilutions of 1:50, would provide an optimal assay signal within a relatively short timeframe.</description><subject>Antibody binding assays</subject><subject>Autoantigens - blood</subject><subject>Autoantigens - immunology</subject><subject>Biological and medical sciences</subject><subject>Biomarkers - blood</subject><subject>Biotechnology</subject><subject>Biotin - immunology</subject><subject>Biotin–GAD65</subject><subject>Blood Chemical Analysis - methods</subject><subject>Diabetes Mellitus, Type 1 - blood</subject><subject>Diabetes Mellitus, Type 1 - diagnosis</subject><subject>Diabetes Mellitus, Type 1 - immunology</subject><subject>Enzyme-Linked Immunosorbent Assay - methods</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Glutamate Decarboxylase - blood</subject><subject>Glutamate Decarboxylase - immunology</subject><subject>Humans</subject><subject>Recombinant Fusion Proteins</subject><subject>Reproducibility of Results</subject><subject>Sensitivity and Specificity</subject><subject>Type 1 diabetes</subject><issn>0168-1656</issn><issn>1873-4863</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2004</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqF0ctu1DAUBmALgegw8Aggb0B0keBLfMmqqlpakCqxgbXlOMeVR4k9xEnF7HgH3pAnwcNEgl1Xlqzv-Pzyj9BrSmpKqPywq3ddSDO4mhHS1ITXhLRP0IZqxatGS_4UbYrTFZVCnqEXOe9Iga2gz9EZFbSVSjUbNFzDAwxpP0KccfL4flhmOwaHrQs97sHZqUs_DoPNgKXA728vr6U4x3aZk41z6FJ_wDZne8BLDvEeHzOF-Pvnr78Q-3KbIt5PJWmIL9Ezb4cMr9Zzi77dfPx69am6-3L7-eryrnKNaOdKcSVtp4lV3Avw0rZScN8RpRVzmveWMGg1COdU08tOAnDWKGBMSc46T_gWvTu9W_Z-XyDPZgzZwTDYCGnJRjHCaEv5o5CqVjNd5BaJE3RTynkCb_ZTGO10MJSYYx9mZ9Y-zLEPQ7gpfZS5N-uCpRuh_ze1FlDA2xXY7OzgJxtdyP85LRXTuriLk4Pybw8BJpNdgOigDxO42fQpPBLlD35aq9g</recordid><startdate>20040701</startdate><enddate>20040701</enddate><creator>Luo, Dong</creator><creator>Rogers, Chad N</creator><creator>Steed, Jordan T</creator><creator>Gilliam, Lisa K</creator><creator>Hampe, Christiane S</creator><creator>Lernmark, Åke</creator><general>Elsevier B.V</general><general>Elsevier</general><scope>IQODW</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>7QO</scope><scope>8FD</scope><scope>FR3</scope><scope>P64</scope><scope>7X8</scope></search><sort><creationdate>20040701</creationdate><title>Development of glutamic acid decarboxylase 65 (GAD65) autoantibody assay using biotin–GAD65 fusion protein</title><author>Luo, Dong ; Rogers, Chad N ; Steed, Jordan T ; Gilliam, Lisa K ; Hampe, Christiane S ; Lernmark, Åke</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c459t-7376ab80a73f5ef6a9653fb07872c83da02e98e5cc74d6b6ee3247e227632bf03</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2004</creationdate><topic>Antibody binding assays</topic><topic>Autoantigens - blood</topic><topic>Autoantigens - immunology</topic><topic>Biological and medical sciences</topic><topic>Biomarkers - blood</topic><topic>Biotechnology</topic><topic>Biotin - immunology</topic><topic>Biotin–GAD65</topic><topic>Blood Chemical Analysis - methods</topic><topic>Diabetes Mellitus, Type 1 - blood</topic><topic>Diabetes Mellitus, Type 1 - diagnosis</topic><topic>Diabetes Mellitus, Type 1 - immunology</topic><topic>Enzyme-Linked Immunosorbent Assay - methods</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>Glutamate Decarboxylase - blood</topic><topic>Glutamate Decarboxylase - immunology</topic><topic>Humans</topic><topic>Recombinant Fusion Proteins</topic><topic>Reproducibility of Results</topic><topic>Sensitivity and Specificity</topic><topic>Type 1 diabetes</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Luo, Dong</creatorcontrib><creatorcontrib>Rogers, Chad N</creatorcontrib><creatorcontrib>Steed, Jordan T</creatorcontrib><creatorcontrib>Gilliam, Lisa K</creatorcontrib><creatorcontrib>Hampe, Christiane S</creatorcontrib><creatorcontrib>Lernmark, Åke</creatorcontrib><collection>Pascal-Francis</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Biotechnology Research Abstracts</collection><collection>Technology Research Database</collection><collection>Engineering Research Database</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>MEDLINE - Academic</collection><jtitle>Journal of biotechnology</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Luo, Dong</au><au>Rogers, Chad N</au><au>Steed, Jordan T</au><au>Gilliam, Lisa K</au><au>Hampe, Christiane S</au><au>Lernmark, Åke</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Development of glutamic acid decarboxylase 65 (GAD65) autoantibody assay using biotin–GAD65 fusion protein</atitle><jtitle>Journal of biotechnology</jtitle><addtitle>J Biotechnol</addtitle><date>2004-07-01</date><risdate>2004</risdate><volume>111</volume><issue>1</issue><spage>97</spage><epage>104</epage><pages>97-104</pages><issn>0168-1656</issn><eissn>1873-4863</eissn><coden>JBITD4</coden><abstract>We evaluated a biotin–glutamic acid decarboxylase 65 (GAD65)-based enzyme-linked immunosorbent assay (B-ELISA) to detect GAD65 autoantibodies (GAD65Ab) in 78 sera from individuals with newly diagnosed type 1 diabetes. The GAD65Ab index of patients with type 1 diabetes (mean value of GAD65Ab index of 1.891) was significantly higher than those in 50 sera from healthy control group (mean value of 0.068). The intra- and inter-assay coefficients of variation (CV) were calculated to be 1.042 and 10.703%, respectively. The specificity of the B-GAD65 ELISA was comparable to the standard radioimmunoassay (RIA) which is routinely used in the laboratory. We describe the optimal conditions of the binding kinetics from each assay-step for the detection of GAD65Ab using the WHO standard serum 97/550 as a model autoantibody serum. We concluded that incubation times of 15, 90, and 90
min for step 1 (pre-incubation of Biotin14–GAD65 with serum), step 2 (binding the Ab/Ag complex to NeutrAvidin plate), and step 3 (incubation with HRPO-anti-human IgG), respectively, along with human serum dilutions of 1:50, would provide an optimal assay signal within a relatively short timeframe.</abstract><cop>Lausanne</cop><cop>Amsterdam</cop><cop>New York, NY</cop><pub>Elsevier B.V</pub><pmid>15196774</pmid><doi>10.1016/j.jbiotec.2004.03.009</doi><tpages>8</tpages></addata></record> |
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| subjects | Antibody binding assays Autoantigens - blood Autoantigens - immunology Biological and medical sciences Biomarkers - blood Biotechnology Biotin - immunology Biotin–GAD65 Blood Chemical Analysis - methods Diabetes Mellitus, Type 1 - blood Diabetes Mellitus, Type 1 - diagnosis Diabetes Mellitus, Type 1 - immunology Enzyme-Linked Immunosorbent Assay - methods Fundamental and applied biological sciences. Psychology Glutamate Decarboxylase - blood Glutamate Decarboxylase - immunology Humans Recombinant Fusion Proteins Reproducibility of Results Sensitivity and Specificity Type 1 diabetes |
| title | Development of glutamic acid decarboxylase 65 (GAD65) autoantibody assay using biotin–GAD65 fusion protein |
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