Purification of IgG Using Gel-Filtration Chromatography

Abstract

In gel filtration, a protein mixture (the mobile phase) is applied to a column of small beads with pores of carefully controlled size (the stationary phase). The movement of the solute is dependent on the flow of the mobile phase, and the Brownian motion of the solute molecules causes their diffusion into and out of the chromatographic bed. Large proteins, above the “exclusion limit” of the gel, cannot enter the pore and, hence, are eluted in the “void volume” of the column (see Note 1). Small proteins enter the pores and are therefore eluted in the “total volume” of the column, and intermediate-size proteins are eluted between the void and total volumes. Proteins are therefore eluted in order of decreasing molecular size. Column matrices are available in a number of fractionation ranges, allowing the user to select the appropriate column for their particular application.