High-performance size-exclusion chromatography of recombinant derived proteins and aggregated species

• Published in: Journal of Chromatography A 1988, Vol.436 (2), p.289-298
• Main Authors: Watson, Eric, Kenney, William C.
• Format: Article
• Language: English
• Subjects: Biological and medical sciences; Biotechnology; Fundamental and applied biological sciences. Psychology; Health. Pharmaceutical industry; high-performance liquid chromatography; Industrial applications and implications. Economical aspects; Other active biomolecules; Production of active biomolecules
• ISSN: 0021-9673
• DOI: 10.1016/S0021-9673(00)94586-2

The chromatographic behavior of some recombinant derived proteins and aggregated species was studied using high-performance size-exclusion chromatography (HPSEC). At neutral pH values, monomeric proteins exhibited non-ideal behavior while aggregated species were not eluted. As the pH was lowered below 5, both aggregated and monomeric species were eluted, with the amount of aggregated species increasing with the decrease in pH. Final elution conditions selected for the simultaneous chromatography of monomeric and aggregated proteins were 0.1 M orthophosphoric acid, pH 2.5. The utility of the system was evaluated by determining the rates of protein degradation at elevated temperatures and comparing the results with those obtained using standard bioassay procedures. The rate of formation of aggregated species was also determined by HPSEC and correspond to the rate of degradation of monomeric protein. The use of HPSEC with low pH eluent provides a rapid means for estimating protein stability under accelarated temperature conditions as well as for determining the existence and formation of aggregated species.