Journal Highlight: Comparison of sample preparation techniques for large-scale proteomics

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  • Published: Jan 30, 2017
  • Author: separationsNOW
  • Channels: Proteomics & Genomics
thumbnail image: Journal Highlight: Comparison of sample preparation techniques for large-scale proteomics

The performances of several common sample preparation techniques for proteomic characterization of HeLa lysates have been compared, proteome sequence coverage increasing to 38% when combining all workflows.

Comparison of sample preparation techniques for large-scale proteomics

Proteomics, 2017, 17, online
Miljan Kuljanin, Dylan Z. Dieters-Castator, David A. Hess, Lynne-Marie Postovit and Gilles A. Lajoie

Abstract: Numerous workflows exist for large-scale bottom-up proteomics, many of which achieve exceptional proteome depth. Herein, we evaluated the performance of several commonly used sample preparation techniques for proteomic characterization of HeLa lysates [unfractionated in-solution digests, SDS-PAGE coupled with in-gel digestion, gel-eluted liquid fraction entrapment electrophoresis (GELFrEE) technology, SCX StageTips and high-/low-pH reversed phase fractionation (HpH)]. HpH fractionation was found to be superior in terms of proteome depth (>8400 proteins detected) and fractionation efficiency compared to other techniques. SCX StageTip fractionation required minimal sample handling and was also a substantial improvement over SDS-PAGE separation and GELFrEE technology. Sequence coverage of the HeLa proteome increased to 38% when combining all workflows, however, total proteins detected improved only slightly to 8710. In summary, HpH fractionation and SCX StageTips are robust techniques and highly suited for complex proteome analysis.

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