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Towards a human proteome atlas: High-throughput generation of mono-specific antibodies for tissue profiling
A great need exists for the systematic generation of specific antibodies to explore the human proteome. Here, we show that antibodies specific to human proteins can be generated in a high-throughput manner involving stringent affinity purification using recombinant protein epitope signature tags (Pr...
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| Published in: | Proteomics (Weinheim) 2005-11, Vol.5 (17), p.4327-4337 |
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| Main Authors: | , , , , , , , , , , , , , , , |
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| cites | cdi_FETCH-LOGICAL-c6052-6be451c86b1b09874227ff3dd6edc59cd80f72854e3feec7cb2b4ce583f60f493 |
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| container_title | Proteomics (Weinheim) |
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| creator | Nilsson, Peter Paavilainen, Linda Larsson, Karin Ödling, Jenny Sundberg, Mårten Andersson, Ann-Catrin Kampf, Caroline Persson, Anja Szigyarto, Cristina Al-Khalili Ottosson, Jenny Björling, Erik Hober, Sophia Wernérus, Henrik Wester, Kenneth Pontén, Fredrik Uhlen, Mathias |
| description | A great need exists for the systematic generation of specific antibodies to explore the human proteome. Here, we show that antibodies specific to human proteins can be generated in a high-throughput manner involving stringent affinity purification using recombinant protein epitope signature tags (PrESTs) as immunogens and affinity-ligands. The specificity of the generated affinity reagents, here called mono-specific antibodies (msAb), were validated with a novel protein microarray assay. The success rate for 464 antibodies generated towards human proteins was more than 90% as judged by the protein array assay. The antibodies were used for parallel profiling of patient biopsies using tissue microarrays generated from 48 human tissues. Comparative analysis with well-characterized monoclonal antibodies showed identical or similar specificity and expression patterns. The results suggest that a comprehensive atlas containing extensive protein expression and subcellular localization data of the human proteome can be generated in an efficient manner with mono-specific antibodies. |
| doi_str_mv | 10.1002/pmic.200500072 |
| format | article |
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Here, we show that antibodies specific to human proteins can be generated in a high-throughput manner involving stringent affinity purification using recombinant protein epitope signature tags (PrESTs) as immunogens and affinity-ligands. The specificity of the generated affinity reagents, here called mono-specific antibodies (msAb), were validated with a novel protein microarray assay. The success rate for 464 antibodies generated towards human proteins was more than 90% as judged by the protein array assay. The antibodies were used for parallel profiling of patient biopsies using tissue microarrays generated from 48 human tissues. Comparative analysis with well-characterized monoclonal antibodies showed identical or similar specificity and expression patterns. The results suggest that a comprehensive atlas containing extensive protein expression and subcellular localization data of the human proteome can be generated in an efficient manner with mono-specific antibodies.</description><identifier>ISSN: 1615-9853</identifier><identifier>ISSN: 1615-9861</identifier><identifier>EISSN: 1615-9861</identifier><identifier>DOI: 10.1002/pmic.200500072</identifier><identifier>PMID: 16237735</identifier><language>eng</language><publisher>Weinheim: Wiley-VCH Verlag</publisher><subject>Analytical, structural and metabolic biochemistry ; Animals ; Antibodies - chemistry ; Antibodies - isolation & purification ; Antibody generation ; Antibody Specificity ; Bioengineering ; Biological and medical sciences ; Bioteknik ; Chromatography, Affinity ; Fundamental and applied biological sciences. Psychology ; Humans ; Immunohistochemistry - methods ; Miscellaneous ; Protein Array Analysis ; Protein microarray ; Proteins ; Proteome - chemistry ; Proteome - immunology ; Proteome - isolation & purification ; Proteome atlas ; Rabbits ; Recombinant Proteins - chemistry ; Recombinant Proteins - immunology ; TECHNOLOGY ; TEKNIKVETENSKAP ; Tissue microarray ; Tissue profiling</subject><ispartof>Proteomics (Weinheim), 2005-11, Vol.5 (17), p.4327-4337</ispartof><rights>Copyright © 2005 WILEY‐VCH Verlag GmbH & Co. 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Here, we show that antibodies specific to human proteins can be generated in a high-throughput manner involving stringent affinity purification using recombinant protein epitope signature tags (PrESTs) as immunogens and affinity-ligands. The specificity of the generated affinity reagents, here called mono-specific antibodies (msAb), were validated with a novel protein microarray assay. The success rate for 464 antibodies generated towards human proteins was more than 90% as judged by the protein array assay. The antibodies were used for parallel profiling of patient biopsies using tissue microarrays generated from 48 human tissues. Comparative analysis with well-characterized monoclonal antibodies showed identical or similar specificity and expression patterns. The results suggest that a comprehensive atlas containing extensive protein expression and subcellular localization data of the human proteome can be generated in an efficient manner with mono-specific antibodies.</description><subject>Analytical, structural and metabolic biochemistry</subject><subject>Animals</subject><subject>Antibodies - chemistry</subject><subject>Antibodies - isolation & purification</subject><subject>Antibody generation</subject><subject>Antibody Specificity</subject><subject>Bioengineering</subject><subject>Biological and medical sciences</subject><subject>Bioteknik</subject><subject>Chromatography, Affinity</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Humans</subject><subject>Immunohistochemistry - methods</subject><subject>Miscellaneous</subject><subject>Protein Array Analysis</subject><subject>Protein microarray</subject><subject>Proteins</subject><subject>Proteome - chemistry</subject><subject>Proteome - immunology</subject><subject>Proteome - isolation & purification</subject><subject>Proteome atlas</subject><subject>Rabbits</subject><subject>Recombinant Proteins - chemistry</subject><subject>Recombinant Proteins - immunology</subject><subject>TECHNOLOGY</subject><subject>TEKNIKVETENSKAP</subject><subject>Tissue microarray</subject><subject>Tissue profiling</subject><issn>1615-9853</issn><issn>1615-9861</issn><issn>1615-9861</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2005</creationdate><recordtype>article</recordtype><recordid>eNqF0ctv0zAYAPAIgdgYXDmCL3CBFNuJH9ltKuwhbYBEB1IvluPYiWkSBztR2X-Pq1Qdt578HX7fy1-SvEZwgSDEn4bOqgWGkEAIGX6SnCKKSFpwip4eYpKdJC9C-A0hYrxgz5MTRHHGWEZOk83KbaWvApCgmTrZg8G7UbtOAzm2MpyDa1s36dh4N9XNMI2g1r32crSuB86AzvUuDYNW1lgFZD_a0lVWB2CcB6MNYdK7isa2tq9fJs-MbIN-tX_PkvvLL6vldXr77epmeXGbKgoJTmmpc4IUpyUqYcFZjjEzJqsqqitFClVxaBjmJNeZ0VoxVeIyV5rwzFBo8iI7Sz7MdcNWD1MpBm876R-Ek1Z8tj8vhPO1mCbBIYU46o_H9WZsBEI55pG_n3lc68-kwyg6G5RuW9lrNwVBOceEZuwoRAzF3SCNcDFD5V0IXpvDCAiK3ZHF7sjicOSY8GZfeSo7XT3y_VUjeLcHMijZGi97ZcOjY5gVjO06F7Pb2lY_HGkrvt_dLP8fIp1zbRj130Ou9BtBWcaI-PX1Sqzz1V2OL9diHf3b2RvphKx9nOf-B4Yog_EbKInBPxdA2z4</recordid><startdate>20051101</startdate><enddate>20051101</enddate><creator>Nilsson, Peter</creator><creator>Paavilainen, Linda</creator><creator>Larsson, Karin</creator><creator>Ödling, Jenny</creator><creator>Sundberg, Mårten</creator><creator>Andersson, Ann-Catrin</creator><creator>Kampf, Caroline</creator><creator>Persson, Anja</creator><creator>Szigyarto, Cristina Al-Khalili</creator><creator>Ottosson, Jenny</creator><creator>Björling, Erik</creator><creator>Hober, Sophia</creator><creator>Wernérus, Henrik</creator><creator>Wester, Kenneth</creator><creator>Pontén, Fredrik</creator><creator>Uhlen, Mathias</creator><general>Wiley-VCH Verlag</general><general>WILEY-VCH Verlag</general><general>WILEY‐VCH Verlag</general><general>Wiley-VCH</general><scope>FBQ</scope><scope>BSCLL</scope><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>RC3</scope><scope>7X8</scope><scope>ADTPV</scope><scope>AOWAS</scope><scope>D8V</scope><scope>DF2</scope></search><sort><creationdate>20051101</creationdate><title>Towards a human proteome atlas: High-throughput generation of mono-specific antibodies for tissue profiling</title><author>Nilsson, Peter ; Paavilainen, Linda ; Larsson, Karin ; Ödling, Jenny ; Sundberg, Mårten ; Andersson, Ann-Catrin ; Kampf, Caroline ; Persson, Anja ; Szigyarto, Cristina Al-Khalili ; Ottosson, Jenny ; Björling, Erik ; Hober, Sophia ; Wernérus, Henrik ; Wester, Kenneth ; Pontén, Fredrik ; Uhlen, Mathias</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c6052-6be451c86b1b09874227ff3dd6edc59cd80f72854e3feec7cb2b4ce583f60f493</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2005</creationdate><topic>Analytical, structural and metabolic biochemistry</topic><topic>Animals</topic><topic>Antibodies - chemistry</topic><topic>Antibodies - isolation & purification</topic><topic>Antibody generation</topic><topic>Antibody Specificity</topic><topic>Bioengineering</topic><topic>Biological and medical sciences</topic><topic>Bioteknik</topic><topic>Chromatography, Affinity</topic><topic>Fundamental and applied biological sciences. 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| ispartof | Proteomics (Weinheim), 2005-11, Vol.5 (17), p.4327-4337 |
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| subjects | Analytical, structural and metabolic biochemistry Animals Antibodies - chemistry Antibodies - isolation & purification Antibody generation Antibody Specificity Bioengineering Biological and medical sciences Bioteknik Chromatography, Affinity Fundamental and applied biological sciences. Psychology Humans Immunohistochemistry - methods Miscellaneous Protein Array Analysis Protein microarray Proteins Proteome - chemistry Proteome - immunology Proteome - isolation & purification Proteome atlas Rabbits Recombinant Proteins - chemistry Recombinant Proteins - immunology TECHNOLOGY TEKNIKVETENSKAP Tissue microarray Tissue profiling |
| title | Towards a human proteome atlas: High-throughput generation of mono-specific antibodies for tissue profiling |
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