Dialysis and Concentration of Protein Solutions

Sarah M. Andrew1, Julie A. Titus2, Louis Zumstein3

1 Chester College, Chester, United Kingdom, 2 National Cancer Institute, Bethesda, Maryland, 3 Introgen Therapeutics, Inc., Houston, Texas
Publication Name:  Current Protocols in Toxicology
Unit Number:  Appendix 3H
DOI:  10.1002/0471140856.txa03hs10
Online Posting Date:  February, 2002
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Abstract

Conventional dialysis separates small molecules from large molecules by allowing diffusion of only the small molecules through selectively permeable membranes. Dialysis is usually used to change the salt (small‐molecule) composition of a macromolecule‐containing solution. The solution to be dialyzed is placed in a sealed dialysis membrane and immersed in a selected buffer; small solute molecules then equilibrate between the sample and the dialysate. Concomitant with the movement of small solutes across the membrane, however, is the movement of solvent in the opposite direction. There are several simple and relatively inexpensive methods for concentrating protein solutions. Dialysis against Aquacide 11A (Calbiochem), which removes water through the dialysis tubing, may be used. After concentration, the solution must be redialyzed into the appropriate buffer. Another method is to use Immersible‐CX Ultrafilters (Millipore) which, when connected to a vacuum, remove everything below their molecular weight cutoff (MWCO). Alternatively, centrifugal concentrators, which are operated with the aid of ordinary laboratory centrifuges may be used.

     
 
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Table of Contents

  • Dialysis
  • Basic Protocol 1: Large‐Volume Dialysis
  • Alternate Protocol 1: Small‐Volume Dialysis
  • Support Protocol 1: Selection and Preparation of Dialysis Membrane
  • Basic Protocol 2: Concentration
  • Figures
     
 
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Materials

Basic Protocol 1: Large‐Volume Dialysis

  Materials
  • Dialysis membrane (see protocol 3)
  • Clamps (Spectra/Por Closures, Spectrum, or equivalent)
  • Macromolecule‐containing sample to be dialyzed
  • Appropriate dialysis buffer

Alternate Protocol 1: Small‐Volume Dialysis

  • 0.5‐ml microcentrifuge tube
  • Pasteur pipet
  • Cork borer

Support Protocol 1: Selection and Preparation of Dialysis Membrane

  Materials
  • Dialysis membrane
  • 10 mM sodium bicarbonate
  • 10 mM Na 2EDTA, pH 8.0
  • 20% to 50% (v/v) ethanol
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Figures

Videos

Literature Cited

Literature Cited
   Craig, L.C. 1967. Techniques for the study of peptides and proteins by dialysis and diffusion. Methods Enzymol. 11:870‐905.
Key References
   Craig. 1967. See above.
  Describes the theory and practice of dialysis and diffusion.
   McPhie, P. 1971. Dialysis. Methods Enzymol. 22:23‐32.
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