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Dialysis and Concentration of Protein Solutions
DIALYSIS 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. This can result in some sample dilution (usually 1 mg/ml, it may be necessary to concentrate the solution. In addition, concentration of protein solutions is often necessary to decrease the volume of a sample; e.g., to load onto a size-exclusion column (APPENDIX 3I) when the volume must be ≤5 ml. 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, Centricon microconcentrators (Amicon), which are available in different MWCO sizes, are very useful for concentrating small amounts of material in small volumes. Follow the instructions from the manufacturers when using these products.

Dialysis and Concentration of Protein Solutions

A.3H.4
Supplement 21 Current Protocols in Immunology

KEY REFERENCES
Craig, L.C. 1967. Techniques for the study of peptides and proteins by dialysis and diffusion. Methods Enzymol. 11:870-905. McPhie, P. 1971. Dialysis. Methods Enzymol. 22:23-32. Describes the theory and practice of dialysis and