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Depleting high-abundant and enriching low-abundant proteins in human serum: An evaluation of sample preparation methods using magnetic nanoparticle, chemical depletion and immunoaffinity techniques.
de Jesus, Jemmyson Romário; da Silva Fernandes, Rafael; de Souza Pessôa, Gustavo; Raimundo, Ivo Milton; Arruda, Marco Aurélio Zezzi.
Afiliação
  • de Jesus JR; Spectrometry, Sample Preparation and Mechanization Group - GEPAM, Institute of Chemistry, University of Campinas - UNICAMP, Campinas, Brazil; National Institute of Science and Technology for Bioanalytics, Institute of Chemistry, University of Campinas - UNICAMP, Campinas, Brazil.
  • da Silva Fernandes R; Group for Instrumentation and Automation in Analytical Chemistry - GIA Institute of Chemistry, University of Campinas - UNICAMP.
  • de Souza Pessôa G; Spectrometry, Sample Preparation and Mechanization Group - GEPAM, Institute of Chemistry, University of Campinas - UNICAMP, Campinas, Brazil; National Institute of Science and Technology for Bioanalytics, Institute of Chemistry, University of Campinas - UNICAMP, Campinas, Brazil.
  • Raimundo IM; Group for Instrumentation and Automation in Analytical Chemistry - GIA Institute of Chemistry, University of Campinas - UNICAMP.
  • Arruda MAZ; Spectrometry, Sample Preparation and Mechanization Group - GEPAM, Institute of Chemistry, University of Campinas - UNICAMP, Campinas, Brazil; National Institute of Science and Technology for Bioanalytics, Institute of Chemistry, University of Campinas - UNICAMP, Campinas, Brazil. Electronic address:
Talanta ; 170: 199-209, 2017 Aug 01.
Article em En | MEDLINE | ID: mdl-28501159
The efficiency of three different depletion methods to remove the most abundant proteins, enriching those human serum proteins with low abundance is checked to make more efficient the search and discovery of biomarkers. These methods utilize magnetic nanoparticles (MNPs), chemical reagents (sequential application of dithiothreitol and acetonitrile, DTT/ACN), and commercial apparatus based on immunoaffinity (ProteoMiner, PM). The comparison between methods shows significant removal of abundant protein, remaining in the supernatant at concentrations of 4.6±0.2, 3.6±0.1, and 3.3±0.2µgµL-1 (n=3) for MNPs, DTT/ACN and PM respectively, from a total protein content of 54µgµL-1. Using GeLC-MS/MS analysis, MNPs depletion shows good efficiency in removing high molecular weight proteins (>80kDa). Due to the synergic effect between the reagents DTT and ACN, DTT/ACN-based depletion offers good performance in the depletion of thiol-rich proteins, such as albumin and transferrin (DTT action), as well as of high molecular weight proteins (ACN action). Furthermore, PM equalization confirms its efficiency in concentrating low-abundant proteins, decreasing the dynamic range of protein levels in human serum. Direct comparison between the treatments reveals 72 proteins identified when using MNP depletion (43 of them exclusively by this method), but only 20 proteins using DTT/ACN (seven exclusively by this method). Additionally, after PM treatment 30 proteins were identified, seven exclusively by this method. Thus, MNPs and DTT/ACN depletion can be simple, quick, cheap, and robust alternatives for immunochemistry-based protein depletion, providing a potential strategy in the search for disease biomarkers.
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Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Proteínas Sanguíneas / Nanopartículas de Magnetita / Fracionamento Químico Tipo de estudo: Evaluation_studies / Prognostic_studies Limite: Humans Idioma: En Revista: Talanta Ano de publicação: 2017 Tipo de documento: Article País de afiliação: Brasil País de publicação: Holanda
Texto completo
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XML
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Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Proteínas Sanguíneas / Nanopartículas de Magnetita / Fracionamento Químico Tipo de estudo: Evaluation_studies / Prognostic_studies Limite: Humans Idioma: En Revista: Talanta Ano de publicação: 2017 Tipo de documento: Article País de afiliação: Brasil País de publicação: Holanda