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Lys-C/Trypsin Tandem-Digestion Protocol for Gel-Free Proteomic Analysis of Colon Biopsies

  • Armin Schniers
  • Yvonne Pasing
  • Terkel HansenEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1959)

Abstract

The protocol presented was specifically optimized for in-depth analysis of the human colon mucosa proteome. After cell lysis in a sodium deoxycholate/urea buffer, a tandem digestion with Lys-C and trypsin was performed. Prior to LC-MS/MS analysis, peptides were TMT-labeled and fractionated by high pH reversed-phase spin columns. This protocol is a powerful, reproducible, sample-saving, and cost-effective option when an in-depth quantitative proteome analysis is desired.

Key words

Colon mucosa proteome Sodium deoxycholate Lys-C TMT labeling High pH reversed-phase fractionation 

Notes

Acknowledgments

We thank Ilona Urbarova and Jack-Ansgar Bruun for fruitful discussions, as well as Prof Jon Florholmen and Rasmus Goll for supplying colon biopsies. Our work was supported by a grant from the North Norway Regional Health Authorities.

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Natural Products and Medicinal Chemistry Research Group, Department of PharmacyUiT—The Arctic University of NorwayTromsøNorway
  2. 2.Tromsø Endocrine Research Group, Department of Clinical MedicineUiT—The Arctic University of NorwayTromsøNorway
  3. 3.Division of Internal MedicineUniversity Hospital of North NorwayTromsøNorway

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