Comparison of different depletion strategies for improving resolution of the human urine proteome
Background: Urine, being an ultrafiltrate of plasma, is a rich source for biomarker discovery. Since potential new disease markers are often present in low concentrations, a prefractionation/enrichment step could be useful in the discovery process. To enhance the detection of low-abundance proteins,...
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Walter de Gruyter ; 2010 |
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Enthalten in: Clinical chemistry and laboratory medicine - Berlin [u.a.] : De Gruyter, 1998, 48(2010), 4 vom: 12. Feb., Seite 531-535 |
Übergeordnetes Werk: |
volume:48 ; year:2010 ; number:4 ; day:12 ; month:02 ; pages:531-535 ; extent:5 |
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DOI / URN: |
10.1515/CCLM.2010.109 |
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NLEJ246730161 |
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10.1515/CCLM.2010.109 doi artikel_Grundlieferung.pp (DE-627)NLEJ246730161 DE-627 ger DE-627 rakwb Comparison of different depletion strategies for improving resolution of the human urine proteome Walter de Gruyter 2010 5 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier Background: Urine, being an ultrafiltrate of plasma, is a rich source for biomarker discovery. Since potential new disease markers are often present in low concentrations, a prefractionation/enrichment step could be useful in the discovery process. To enhance the detection of low-abundance proteins, three immuno-affinity depletion approaches were evaluated. Methods: To remove the most abundant proteins from a human urine sample, GenWay™ Spin IgY-12 kit, HPLC Agilent Hu-PL7 and a home-made column vs. human serum albumin [immuno-affinity column (IAC)] were compared. Quantification of total proteins, 2-D gel electrophoresis (2-DE), Progenesis gel images analysis and mass spectrometric proteins identification were applied to evaluate these strategies. Results: Reproducibility of depletion columns, by estimating protein content of unbound fractions, were: 343±20.0 μg, 5.8%; 186.3±13.3 μg, 7.2%; 292±20.6 μg, 8.8% [mean± standard deviation (SD), CV%], for GenWay™, Agilent and IAC methods, respectively. To isolate urinary protein after depletion, ethanol precipitation provided the highest recovery (80%). Applying 2-DE and Progenesis analysis, the number of spots visualized on the gels was 468±21, 331±7, 368±22 and 304±7 (mean±SD) for GenWay™, Agilent, IAC, and the undepleted urine pool sample, respectively, with a significant difference p<0.001 compared to the GenWay procedure. Conclusions: The sequential procedure of urine samples using multi-protein immuno-affinity depletion represents a valid tool for simplifying 2-DE analysis of the urine proteome. Particularly, the GenWay™ kit followed by ethanol precipitation was found to be the most efficient method for exploring the urine proteome. Clin Chem Lab Med 2010;48:531–5. Walter de Gruyter Online Zeitschriften biomarkers proteins depletion proteomics urine Magagnotti, Cinzia oth Fermo, Isabella oth Carletti, Rose Mary oth Ferrari, Maurizio oth Bachi, Angela oth Enthalten in Clinical chemistry and laboratory medicine Berlin [u.a.] : De Gruyter, 1998 48(2010), 4 vom: 12. Feb., Seite 531-535 (DE-627)NLEJ248235222 (DE-600)1492732-9 1437-4331 nnns volume:48 year:2010 number:4 day:12 month:02 pages:531-535 extent:5 https://doi.org/10.1515/CCLM.2010.109 Deutschlandweit zugänglich GBV_USEFLAG_U ZDB-1-DGR GBV_NL_ARTICLE AR 48 2010 4 12 02 531-535 5 |
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10.1515/CCLM.2010.109 doi artikel_Grundlieferung.pp (DE-627)NLEJ246730161 DE-627 ger DE-627 rakwb Comparison of different depletion strategies for improving resolution of the human urine proteome Walter de Gruyter 2010 5 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier Background: Urine, being an ultrafiltrate of plasma, is a rich source for biomarker discovery. Since potential new disease markers are often present in low concentrations, a prefractionation/enrichment step could be useful in the discovery process. To enhance the detection of low-abundance proteins, three immuno-affinity depletion approaches were evaluated. Methods: To remove the most abundant proteins from a human urine sample, GenWay™ Spin IgY-12 kit, HPLC Agilent Hu-PL7 and a home-made column vs. human serum albumin [immuno-affinity column (IAC)] were compared. Quantification of total proteins, 2-D gel electrophoresis (2-DE), Progenesis gel images analysis and mass spectrometric proteins identification were applied to evaluate these strategies. Results: Reproducibility of depletion columns, by estimating protein content of unbound fractions, were: 343±20.0 μg, 5.8%; 186.3±13.3 μg, 7.2%; 292±20.6 μg, 8.8% [mean± standard deviation (SD), CV%], for GenWay™, Agilent and IAC methods, respectively. To isolate urinary protein after depletion, ethanol precipitation provided the highest recovery (80%). Applying 2-DE and Progenesis analysis, the number of spots visualized on the gels was 468±21, 331±7, 368±22 and 304±7 (mean±SD) for GenWay™, Agilent, IAC, and the undepleted urine pool sample, respectively, with a significant difference p<0.001 compared to the GenWay procedure. Conclusions: The sequential procedure of urine samples using multi-protein immuno-affinity depletion represents a valid tool for simplifying 2-DE analysis of the urine proteome. Particularly, the GenWay™ kit followed by ethanol precipitation was found to be the most efficient method for exploring the urine proteome. Clin Chem Lab Med 2010;48:531–5. Walter de Gruyter Online Zeitschriften biomarkers proteins depletion proteomics urine Magagnotti, Cinzia oth Fermo, Isabella oth Carletti, Rose Mary oth Ferrari, Maurizio oth Bachi, Angela oth Enthalten in Clinical chemistry and laboratory medicine Berlin [u.a.] : De Gruyter, 1998 48(2010), 4 vom: 12. Feb., Seite 531-535 (DE-627)NLEJ248235222 (DE-600)1492732-9 1437-4331 nnns volume:48 year:2010 number:4 day:12 month:02 pages:531-535 extent:5 https://doi.org/10.1515/CCLM.2010.109 Deutschlandweit zugänglich GBV_USEFLAG_U ZDB-1-DGR GBV_NL_ARTICLE AR 48 2010 4 12 02 531-535 5 |
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10.1515/CCLM.2010.109 doi artikel_Grundlieferung.pp (DE-627)NLEJ246730161 DE-627 ger DE-627 rakwb Comparison of different depletion strategies for improving resolution of the human urine proteome Walter de Gruyter 2010 5 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier Background: Urine, being an ultrafiltrate of plasma, is a rich source for biomarker discovery. Since potential new disease markers are often present in low concentrations, a prefractionation/enrichment step could be useful in the discovery process. To enhance the detection of low-abundance proteins, three immuno-affinity depletion approaches were evaluated. Methods: To remove the most abundant proteins from a human urine sample, GenWay™ Spin IgY-12 kit, HPLC Agilent Hu-PL7 and a home-made column vs. human serum albumin [immuno-affinity column (IAC)] were compared. Quantification of total proteins, 2-D gel electrophoresis (2-DE), Progenesis gel images analysis and mass spectrometric proteins identification were applied to evaluate these strategies. Results: Reproducibility of depletion columns, by estimating protein content of unbound fractions, were: 343±20.0 μg, 5.8%; 186.3±13.3 μg, 7.2%; 292±20.6 μg, 8.8% [mean± standard deviation (SD), CV%], for GenWay™, Agilent and IAC methods, respectively. To isolate urinary protein after depletion, ethanol precipitation provided the highest recovery (80%). Applying 2-DE and Progenesis analysis, the number of spots visualized on the gels was 468±21, 331±7, 368±22 and 304±7 (mean±SD) for GenWay™, Agilent, IAC, and the undepleted urine pool sample, respectively, with a significant difference p<0.001 compared to the GenWay procedure. Conclusions: The sequential procedure of urine samples using multi-protein immuno-affinity depletion represents a valid tool for simplifying 2-DE analysis of the urine proteome. Particularly, the GenWay™ kit followed by ethanol precipitation was found to be the most efficient method for exploring the urine proteome. Clin Chem Lab Med 2010;48:531–5. Walter de Gruyter Online Zeitschriften biomarkers proteins depletion proteomics urine Magagnotti, Cinzia oth Fermo, Isabella oth Carletti, Rose Mary oth Ferrari, Maurizio oth Bachi, Angela oth Enthalten in Clinical chemistry and laboratory medicine Berlin [u.a.] : De Gruyter, 1998 48(2010), 4 vom: 12. Feb., Seite 531-535 (DE-627)NLEJ248235222 (DE-600)1492732-9 1437-4331 nnns volume:48 year:2010 number:4 day:12 month:02 pages:531-535 extent:5 https://doi.org/10.1515/CCLM.2010.109 Deutschlandweit zugänglich GBV_USEFLAG_U ZDB-1-DGR GBV_NL_ARTICLE AR 48 2010 4 12 02 531-535 5 |
allfieldsGer |
10.1515/CCLM.2010.109 doi artikel_Grundlieferung.pp (DE-627)NLEJ246730161 DE-627 ger DE-627 rakwb Comparison of different depletion strategies for improving resolution of the human urine proteome Walter de Gruyter 2010 5 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier Background: Urine, being an ultrafiltrate of plasma, is a rich source for biomarker discovery. Since potential new disease markers are often present in low concentrations, a prefractionation/enrichment step could be useful in the discovery process. To enhance the detection of low-abundance proteins, three immuno-affinity depletion approaches were evaluated. Methods: To remove the most abundant proteins from a human urine sample, GenWay™ Spin IgY-12 kit, HPLC Agilent Hu-PL7 and a home-made column vs. human serum albumin [immuno-affinity column (IAC)] were compared. Quantification of total proteins, 2-D gel electrophoresis (2-DE), Progenesis gel images analysis and mass spectrometric proteins identification were applied to evaluate these strategies. Results: Reproducibility of depletion columns, by estimating protein content of unbound fractions, were: 343±20.0 μg, 5.8%; 186.3±13.3 μg, 7.2%; 292±20.6 μg, 8.8% [mean± standard deviation (SD), CV%], for GenWay™, Agilent and IAC methods, respectively. To isolate urinary protein after depletion, ethanol precipitation provided the highest recovery (80%). Applying 2-DE and Progenesis analysis, the number of spots visualized on the gels was 468±21, 331±7, 368±22 and 304±7 (mean±SD) for GenWay™, Agilent, IAC, and the undepleted urine pool sample, respectively, with a significant difference p<0.001 compared to the GenWay procedure. Conclusions: The sequential procedure of urine samples using multi-protein immuno-affinity depletion represents a valid tool for simplifying 2-DE analysis of the urine proteome. Particularly, the GenWay™ kit followed by ethanol precipitation was found to be the most efficient method for exploring the urine proteome. Clin Chem Lab Med 2010;48:531–5. Walter de Gruyter Online Zeitschriften biomarkers proteins depletion proteomics urine Magagnotti, Cinzia oth Fermo, Isabella oth Carletti, Rose Mary oth Ferrari, Maurizio oth Bachi, Angela oth Enthalten in Clinical chemistry and laboratory medicine Berlin [u.a.] : De Gruyter, 1998 48(2010), 4 vom: 12. Feb., Seite 531-535 (DE-627)NLEJ248235222 (DE-600)1492732-9 1437-4331 nnns volume:48 year:2010 number:4 day:12 month:02 pages:531-535 extent:5 https://doi.org/10.1515/CCLM.2010.109 Deutschlandweit zugänglich GBV_USEFLAG_U ZDB-1-DGR GBV_NL_ARTICLE AR 48 2010 4 12 02 531-535 5 |
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10.1515/CCLM.2010.109 doi artikel_Grundlieferung.pp (DE-627)NLEJ246730161 DE-627 ger DE-627 rakwb Comparison of different depletion strategies for improving resolution of the human urine proteome Walter de Gruyter 2010 5 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier Background: Urine, being an ultrafiltrate of plasma, is a rich source for biomarker discovery. Since potential new disease markers are often present in low concentrations, a prefractionation/enrichment step could be useful in the discovery process. To enhance the detection of low-abundance proteins, three immuno-affinity depletion approaches were evaluated. Methods: To remove the most abundant proteins from a human urine sample, GenWay™ Spin IgY-12 kit, HPLC Agilent Hu-PL7 and a home-made column vs. human serum albumin [immuno-affinity column (IAC)] were compared. Quantification of total proteins, 2-D gel electrophoresis (2-DE), Progenesis gel images analysis and mass spectrometric proteins identification were applied to evaluate these strategies. Results: Reproducibility of depletion columns, by estimating protein content of unbound fractions, were: 343±20.0 μg, 5.8%; 186.3±13.3 μg, 7.2%; 292±20.6 μg, 8.8% [mean± standard deviation (SD), CV%], for GenWay™, Agilent and IAC methods, respectively. To isolate urinary protein after depletion, ethanol precipitation provided the highest recovery (80%). Applying 2-DE and Progenesis analysis, the number of spots visualized on the gels was 468±21, 331±7, 368±22 and 304±7 (mean±SD) for GenWay™, Agilent, IAC, and the undepleted urine pool sample, respectively, with a significant difference p<0.001 compared to the GenWay procedure. Conclusions: The sequential procedure of urine samples using multi-protein immuno-affinity depletion represents a valid tool for simplifying 2-DE analysis of the urine proteome. Particularly, the GenWay™ kit followed by ethanol precipitation was found to be the most efficient method for exploring the urine proteome. Clin Chem Lab Med 2010;48:531–5. Walter de Gruyter Online Zeitschriften biomarkers proteins depletion proteomics urine Magagnotti, Cinzia oth Fermo, Isabella oth Carletti, Rose Mary oth Ferrari, Maurizio oth Bachi, Angela oth Enthalten in Clinical chemistry and laboratory medicine Berlin [u.a.] : De Gruyter, 1998 48(2010), 4 vom: 12. Feb., Seite 531-535 (DE-627)NLEJ248235222 (DE-600)1492732-9 1437-4331 nnns volume:48 year:2010 number:4 day:12 month:02 pages:531-535 extent:5 https://doi.org/10.1515/CCLM.2010.109 Deutschlandweit zugänglich GBV_USEFLAG_U ZDB-1-DGR GBV_NL_ARTICLE AR 48 2010 4 12 02 531-535 5 |
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Comparison of different depletion strategies for improving resolution of the human urine proteome |
abstract |
Background: Urine, being an ultrafiltrate of plasma, is a rich source for biomarker discovery. Since potential new disease markers are often present in low concentrations, a prefractionation/enrichment step could be useful in the discovery process. To enhance the detection of low-abundance proteins, three immuno-affinity depletion approaches were evaluated. Methods: To remove the most abundant proteins from a human urine sample, GenWay™ Spin IgY-12 kit, HPLC Agilent Hu-PL7 and a home-made column vs. human serum albumin [immuno-affinity column (IAC)] were compared. Quantification of total proteins, 2-D gel electrophoresis (2-DE), Progenesis gel images analysis and mass spectrometric proteins identification were applied to evaluate these strategies. Results: Reproducibility of depletion columns, by estimating protein content of unbound fractions, were: 343±20.0 μg, 5.8%; 186.3±13.3 μg, 7.2%; 292±20.6 μg, 8.8% [mean± standard deviation (SD), CV%], for GenWay™, Agilent and IAC methods, respectively. To isolate urinary protein after depletion, ethanol precipitation provided the highest recovery (80%). Applying 2-DE and Progenesis analysis, the number of spots visualized on the gels was 468±21, 331±7, 368±22 and 304±7 (mean±SD) for GenWay™, Agilent, IAC, and the undepleted urine pool sample, respectively, with a significant difference p<0.001 compared to the GenWay procedure. Conclusions: The sequential procedure of urine samples using multi-protein immuno-affinity depletion represents a valid tool for simplifying 2-DE analysis of the urine proteome. Particularly, the GenWay™ kit followed by ethanol precipitation was found to be the most efficient method for exploring the urine proteome. Clin Chem Lab Med 2010;48:531–5. |
abstractGer |
Background: Urine, being an ultrafiltrate of plasma, is a rich source for biomarker discovery. Since potential new disease markers are often present in low concentrations, a prefractionation/enrichment step could be useful in the discovery process. To enhance the detection of low-abundance proteins, three immuno-affinity depletion approaches were evaluated. Methods: To remove the most abundant proteins from a human urine sample, GenWay™ Spin IgY-12 kit, HPLC Agilent Hu-PL7 and a home-made column vs. human serum albumin [immuno-affinity column (IAC)] were compared. Quantification of total proteins, 2-D gel electrophoresis (2-DE), Progenesis gel images analysis and mass spectrometric proteins identification were applied to evaluate these strategies. Results: Reproducibility of depletion columns, by estimating protein content of unbound fractions, were: 343±20.0 μg, 5.8%; 186.3±13.3 μg, 7.2%; 292±20.6 μg, 8.8% [mean± standard deviation (SD), CV%], for GenWay™, Agilent and IAC methods, respectively. To isolate urinary protein after depletion, ethanol precipitation provided the highest recovery (80%). Applying 2-DE and Progenesis analysis, the number of spots visualized on the gels was 468±21, 331±7, 368±22 and 304±7 (mean±SD) for GenWay™, Agilent, IAC, and the undepleted urine pool sample, respectively, with a significant difference p<0.001 compared to the GenWay procedure. Conclusions: The sequential procedure of urine samples using multi-protein immuno-affinity depletion represents a valid tool for simplifying 2-DE analysis of the urine proteome. Particularly, the GenWay™ kit followed by ethanol precipitation was found to be the most efficient method for exploring the urine proteome. Clin Chem Lab Med 2010;48:531–5. |
abstract_unstemmed |
Background: Urine, being an ultrafiltrate of plasma, is a rich source for biomarker discovery. Since potential new disease markers are often present in low concentrations, a prefractionation/enrichment step could be useful in the discovery process. To enhance the detection of low-abundance proteins, three immuno-affinity depletion approaches were evaluated. Methods: To remove the most abundant proteins from a human urine sample, GenWay™ Spin IgY-12 kit, HPLC Agilent Hu-PL7 and a home-made column vs. human serum albumin [immuno-affinity column (IAC)] were compared. Quantification of total proteins, 2-D gel electrophoresis (2-DE), Progenesis gel images analysis and mass spectrometric proteins identification were applied to evaluate these strategies. Results: Reproducibility of depletion columns, by estimating protein content of unbound fractions, were: 343±20.0 μg, 5.8%; 186.3±13.3 μg, 7.2%; 292±20.6 μg, 8.8% [mean± standard deviation (SD), CV%], for GenWay™, Agilent and IAC methods, respectively. To isolate urinary protein after depletion, ethanol precipitation provided the highest recovery (80%). Applying 2-DE and Progenesis analysis, the number of spots visualized on the gels was 468±21, 331±7, 368±22 and 304±7 (mean±SD) for GenWay™, Agilent, IAC, and the undepleted urine pool sample, respectively, with a significant difference p<0.001 compared to the GenWay procedure. Conclusions: The sequential procedure of urine samples using multi-protein immuno-affinity depletion represents a valid tool for simplifying 2-DE analysis of the urine proteome. Particularly, the GenWay™ kit followed by ethanol precipitation was found to be the most efficient method for exploring the urine proteome. Clin Chem Lab Med 2010;48:531–5. |
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title_short |
Comparison of different depletion strategies for improving resolution of the human urine proteome |
url |
https://doi.org/10.1515/CCLM.2010.109 |
remote_bool |
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author2 |
Magagnotti, Cinzia Fermo, Isabella Carletti, Rose Mary Ferrari, Maurizio Bachi, Angela |
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Magagnotti, Cinzia Fermo, Isabella Carletti, Rose Mary Ferrari, Maurizio Bachi, Angela |
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NLEJ248235222 |
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10.1515/CCLM.2010.109 |
up_date |
2024-07-06T09:12:44.226Z |
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