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SP2: Rapid and Automatable Contaminant Removal from Peptide Samples for Proteomic Analyses. J Proteome Res 2019 Apr 05;18(4):1644-1656

Date

02/24/2019

Pubmed ID

30795648

Pubmed Central ID

PMC6571012

DOI

10.1021/acs.jproteome.8b00916

Scopus ID

2-s2.0-85062444095 (requires institutional sign-in at Scopus site)   30 Citations

Abstract

Peptide cleanup is essential for the removal of contaminating substances that may be introduced during sample preparation steps in bottom-up proteomic workflows. Recent studies have described benefits of carboxylate-modified paramagnetic particles over traditional reversed-phase methods for detergent and polymer removal, but challenges with reproducibility have limited the widespread implementation of this approach among laboratories. To overcome these challenges, the current study systematically evaluated key experimental parameters regarding the use of carboxylate-modified paramagnetic particles and determined those that are critical for maximum performance and peptide recovery and those for which the protocol is tolerant to deviation. These results supported the development of a detailed, easy-to-use standard operating protocol, termed SP2, which can be applied to remove detergents and polymers from peptide samples while concentrating the sample in solvent that is directly compatible with typical LC-MS workflows. We demonstrate that SP2 can be applied to phosphopeptides and glycopeptides and that the approach is compatible with robotic liquid handling for automated sample processing. Altogether, the results of this study and accompanying detailed operating protocols for both manual and automated processing are expected to facilitate reproducible implementation of SP2 for various proteomics applications and will especially benefit core or shared resource facilities where unknown or unexpected contaminants may be particularly problematic.

Author List

Waas M, Pereckas M, Jones Lipinski RA, Ashwood C, Gundry RL

Author

Rachel Jones Lipinski Research Scientist I in the Biochemistry department at Medical College of Wisconsin




MESH terms used to index this publication - Major topics in bold

Chromatography, Liquid
Detergents
HEK293 Cells
Humans
Peptides
Polymers
Proteome
Proteomics
Tandem Mass Spectrometry