Back
Reproducible Tissue Homogenization and Protein Extraction for Quantitative Proteomics Using MicroPestle-Assisted Pressure-Cycling Technology.
Journal article

Reproducible Tissue Homogenization and Protein Extraction for Quantitative Proteomics Using MicroPestle-Assisted Pressure-Cycling Technology.

  • Shao S Department of Biology, Institute of Molecular Systems Biology, ETH Zurich , Zurich, CH-8057 Switzerland.
  • Guo T Department of Biology, Institute of Molecular Systems Biology, ETH Zurich , Zurich, CH-8057 Switzerland.
  • Gross V Pressure BioSciences, Inc. , South Easton, Massachusetts, 02375 United States.
  • Lazarev A Pressure BioSciences, Inc. , South Easton, Massachusetts, 02375 United States.
  • Koh CC Department of Biology, Institute of Molecular Systems Biology, ETH Zurich , Zurich, CH-8057 Switzerland.
  • Gillessen S Department of Biology, Institute of Molecular Systems Biology, ETH Zurich , Zurich, CH-8057 Switzerland.
  • Joerger M
  • Jochum W
  • Aebersold R
Show more…
  • 2016-04-22
Published in:
  • Journal of proteome research. - 2016
PCT-MicroCap
PCT-MicroPestle
SWATH
mass spectrometry
pressure-cycling technology
Animals
Biopsy
Mice
Pressure
Proteins
Proteomics
Reproducibility of Results
Technology
Workflow
English The reproducible and efficient extraction of proteins from biopsy samples for quantitative analysis is a critical step in biomarker and translational research. Recently, we described a method consisting of pressure-cycling technology (PCT) and sequential windowed acquisition of all theoretical fragment ions-mass spectrometry (SWATH-MS) for the rapid quantification of thousands of proteins from biopsy-size tissue samples. As an improvement of the method, we have incorporated the PCT-MicroPestle into the PCT-SWATH workflow. The PCT-MicroPestle is a novel, miniaturized, disposable mechanical tissue homogenizer that fits directly into the microTube sample container. We optimized the pressure-cycling conditions for tissue lysis with the PCT-MicroPestle and benchmarked the performance of the system against the conventional PCT-MicroCap method using mouse liver, heart, brain, and human kidney tissues as test samples. The data indicate that the digestion of the PCT-MicroPestle-extracted proteins yielded 20-40% more MS-ready peptide mass from all tissues tested with a comparable reproducibility when compared to the conventional PCT method. Subsequent SWATH-MS analysis identified a higher number of biologically informative proteins from a given sample. In conclusion, we have developed a new device that can be seamlessly integrated into the PCT-SWATH workflow, leading to increased sample throughput and improved reproducibility at both the protein extraction and proteomic analysis levels when applied to the quantitative proteomic analysis of biopsy-level samples.
Language
  • English
Open access status
closed
Identifiers
Persistent URL
https://sonar.rero.ch/global/documents/22671
Statistics
Document views: 14 File downloads: