NONSPECIFIC INTERACTION OF PROTEOGLYCANS WITH CHROMATOGRAPHY MEDIA AND SURFACES - EFFECT OF THIS INTERACTION ON THE ISOLATION EFFICIENCIES

Nonspecific adsorption of proteoglycans to chromatography media and surfaces is demonstrated. This adsorption is highly dependent on the nature of the chromatography media and the precise buffer conditions. For a given buffer the amount of adsorption decreases as the pH of the buffer is increased. It is also highly dependent on buffer concentration and increases as the buffer concentration is increased. The effect of salts such as LiCl, NaCl, KCl, and MgCl2 was generally small and complex so that the presence of the salt both increased and decreased the amount of adsorption depending on the buffer conditions. In contrast, the effect due to the presence of guanidine hydrochloride (Gdn-HCl) was relatively large and complex. At low Gdn-HCl concentrations there generally was a large increase in the amount of adsorption, reaching a maximum at approximately 0.5 m Gdn-HCl and decreasing with further increases in Gdn-HCl concentration. Detergents such as 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (Chaps) and sodium dodecylsulfate generally reduced the amount of nonspecific adsorption, although in the presence of both the detergent and Gdn-HCl, the effect due to Gdn-HCl predominated. In commonly used buffers such as 0.5 m sodium acetate (NaOAc), pH 7.0 (buffer F), and 4 m Gdn-HCl in 0.05 m NaOAc, pH 5.8 (buffer D), adsorption to surfaces and chromatography media such as Sepharose CL-2B, cellulose, and controlled pore glass (CPG) is highly significant and it is particularly large for cellulose and CPG. Several buffers are described such as 0.05 m Tris-HCl, pH 9.0 (buffer A), and 0.05 m Tris-HCl + 0.5% (w/v) Chaps (buffer C) in which the nonspecific adsorption of proteoglycans to surfaces and chromatography media is minimized. In use of the above buffers for elution of proteoglycans on columns containing chromatography media such as Sepharose CL-2B, Sephadex G-50, cellulose, and CPG, it was shown that there is a close correlation between the amount of nonspecific adsorption and percentage recoveries, which were very good in buffers A and C (84-95%) and poor in buffers D and F (11-70%). © 1990.