Abstract

© Springer Science+Business Media New York 2015. All rights are reserved. All researchers immersed in the world of recombinant protein production are in agreement that often the production and purification process of a protein can become a nightmare due to an unexpected behavior of the protein at different protocol stages. Once the protein is purified, scientists know that they still cannot relax. There is a decisive last step missing: performing a protein dialysis in a suitable buffer for subsequent experimental trials. Here is when we can find proteins that precipitate during dialysis by buffer-related factors (ionic strength, pH, etc.), which are intrinsic to each protein and are difficult to predict. How can we find the buffer in which a protein is more stable and with less tendency to precipitate? In this chapter we go over possible factors affecting the protein precipitation tendency during the dialysis process and describe a general dialysis protocol with tricks to reduce protein aggregation. Furthermore, we propose a fast method to detect the most appropriate buffer for the stability of a particular protein, performing microdialysis on a battery of different buffers to measure afterwards precipitation by a colorimetric method, and thus being able to choose the most suitable buffer for the dialysis of a given protein.
Original languageEnglish
Title of host publicationInsoluble Proteins: Methods and Protocols
Pages321-330
Number of pages9
DOIs
Publication statusPublished - 1 Dec 2014

Keywords

  • Dialysis
  • Microdialysis
  • Protein aggregation
  • Protein precipitation
  • Protein stability

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