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All cats are gray in the dark: enrichment/depletion approaches for biomarker discovery on felis catus plasma


Carcangiu, L; Pisanu, S; Tore, S; Addis, M F; Zini, Eric; Uzzau, S; Pagnozzi, D (2018). All cats are gray in the dark: enrichment/depletion approaches for biomarker discovery on felis catus plasma. Proteomics, 18(20):e1800191.

Abstract

In veterinary medicine, assay performance is often affected by the lack of species-specific diagnostic tools. Reliable biomarkers might be identified by investigating biological fluids of the species of interest, but protein sequence databases are often incomplete and human-specific devices for reducing sample complexity might fail when applied to animal plasma. Here, seven commercial methods based on different capturing agents (anti-human antibodies, affinity ligands, mixture of antibodies and ligands, and combinatorial peptide ligand libraries) are applied to cat plasma and evaluated in terms of yield, identified proteins/ peptides, and relative abundance by high-resolution shotgun proteomics and label-free quantitation. As a result, anti-human antibody-based methods are unsatisfactory. Most fail in reducing albumin and immunoglobulins, and some lead to a substantial removal of other highly abundant proteins, probably because of nonspecific interactions. A protein A/dye ligand-based method is efficient in reducing immunoglobulins, fibrinogen, and apolipoprotein A1 and A2, but not albumin, and protein identifications do not increase. Only peptide ligand libraries flatten the dynamic range, and increased protein identification (59.0%). Albumin and immunoglobulins are successfully depleted (60.7% and 35.9%, respectively). Although further studies will be required for reinforcing our observations, this work can provide a useful guide for cat plasma pretreatment in biomarker discovery studies.

Abstract

In veterinary medicine, assay performance is often affected by the lack of species-specific diagnostic tools. Reliable biomarkers might be identified by investigating biological fluids of the species of interest, but protein sequence databases are often incomplete and human-specific devices for reducing sample complexity might fail when applied to animal plasma. Here, seven commercial methods based on different capturing agents (anti-human antibodies, affinity ligands, mixture of antibodies and ligands, and combinatorial peptide ligand libraries) are applied to cat plasma and evaluated in terms of yield, identified proteins/ peptides, and relative abundance by high-resolution shotgun proteomics and label-free quantitation. As a result, anti-human antibody-based methods are unsatisfactory. Most fail in reducing albumin and immunoglobulins, and some lead to a substantial removal of other highly abundant proteins, probably because of nonspecific interactions. A protein A/dye ligand-based method is efficient in reducing immunoglobulins, fibrinogen, and apolipoprotein A1 and A2, but not albumin, and protein identifications do not increase. Only peptide ligand libraries flatten the dynamic range, and increased protein identification (59.0%). Albumin and immunoglobulins are successfully depleted (60.7% and 35.9%, respectively). Although further studies will be required for reinforcing our observations, this work can provide a useful guide for cat plasma pretreatment in biomarker discovery studies.

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Additional indexing

Item Type:Journal Article, refereed, original work
Communities & Collections:05 Vetsuisse Faculty > Veterinary Clinic > Department of Small Animals
Dewey Decimal Classification:570 Life sciences; biology
630 Agriculture
Uncontrolled Keywords:biomarker discovery; depletion; enrichment; plasma; shotgun proteomics
Language:English
Date:14 September 2018
Deposited On:07 Dec 2018 15:24
Last Modified:01 Oct 2019 11:37
Publisher:Wiley-Blackwell Publishing, Inc.
ISSN:1615-9853
Additional Information:This is the peer reviewed version of the following article: L. Carcangiu, S. Pisanu, S. Tore, M. F. Addis, E. Zini, S. Uzzau, D. Pagnozzi, Proteomics 2018, 18, 1800191, which has been published in final form at https://doi.org/10.1002/pmic.201800191. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Use of Self-Archived Versions. (http://www.wileyauthors.com/self-archiving)
OA Status:Green
Publisher DOI:https://doi.org/10.1002/pmic.201800191
PubMed ID:30216667

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