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Solubilization of Proteins: The Importance of Lysis Buffer Choice

  • Mandy Peach
  • Noelle Marsh
  • Ewa I. Miskiewicz
  • Daniel J. MacPhee
Part of the Methods in Molecular Biology book series (MIMB, volume 1312)

Abstract

The efficient extraction of proteins of interest from cells and tissues is not always straightforward. Here we demonstrate the differences in extraction of the focal adhesion protein Kindlin-2 from choriocarcinoma cells using NP-40 and RIPA lysis buffer. Furthermore, we demonstrate the use of a more denaturing urea/thiourea lysis buffer for solubilization, by comparing its effectiveness for solubilization of small heat-shock proteins from smooth muscle with the often utilized RIPA lysis buffer. Overall, the results demonstrate the importance of establishing the optimal lysis buffer for specific protein solubilization within the experimental workflow.

Key words

Lysis buffer NP-40 lysis buffer RIPA lysis buffer Urea/thiourea lysis buffer Protein extraction Protein solubilization Small heat-shock proteins Focal adhesion proteins 

Notes

Acknowledgement

This work was supported by a Natural Sciences and Engineering Research Council Discovery Grant (#250218), an Establishment Grant from the Saskatchewan Health Research Foundation (SHRF; #2695), and a regional partnership program grant from SHRF (#2776) and the Canadian Institutes of Health Research (#ROP-101051) to D.J.M. M.P. and N.M. were holders of Alexander Graham Bell NSERC postgraduate fellowships.

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Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  • Mandy Peach
    • 1
  • Noelle Marsh
    • 1
  • Ewa I. Miskiewicz
    • 2
  • Daniel J. MacPhee
    • 2
  1. 1.Division of BioMedical SciencesHealth Sciences CentreSt. John’sCanada
  2. 2.Department of Veterinary Biomedical Sciences, One Reproductive Health Research Group, Western College of Veterinary MedicineUniversity of SaskatchewanSaskatoonCanada

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