Multiple functions of caprylic acid-induced impurity precipitation for process intensification in monoclonal antibody purification.

New emerging technologies delivering benefits in terms of process robustness and economy are an inevitable prerequisite for monoclonal antibody purification processes intensification. Caprylic acid was proven as an effective precipitating agent enabling efficient precipitaton of product- and process-related impurities while leaving the antibody in solution. This purification step at mild acidic pH was therefore introduced in generic antibody platform approaches after Protein A capture and evaluated for its impact regarding process robustness and antibody stability.

Reducing sample complexity of polyclonal human autoantibodies by chromatofocusing.

Chromatofocusing was performed in order to separate a polyclonal antigen-specific mixture of human immunoglobulins (IgGs) that would then allow for further analyses of as few different IgGs as possible. Because polyclonal IgGs only differ by amino acid sequence and possible post-translational modifications but not by molecular weight, we chose chromatofocusing for protein separation by different isoelectric points. We isolated antigen-specific IgGs from commercially available intravenous immunoglobulins (IVIG) using a combination of affinity- and size exclusion-chromatography and in order to reduce the complexity of the starting material IVIG was then replaced by single-donor plasmapheresis material.

Investigation of salt properties with electro-acoustic measurements and their effect on dynamic binding capacity in hydrophobic interaction chromatography.

The pH dependence in hydrophobic interaction chromatography (HIC) is usually discussed exclusively in terms of protein dependence and there are no clear defined trends. Many of the deviations from an ideal solution are caused solely by the high salt concentration, as protein concentration is usually negligible. So pH dependency in hydrophobic interaction chromatography could also be the result of pH dependent changes of ion properties from the salt solution.

Fast determination of conditions for maximum dynamic capacity in cation-exchange chromatography of human monoclonal antibodies.

Dynamic binding capacity (DBC) measurements of cation-exchange resins were performed with two human monoclonal antibodies. DBC showed a pH dependent maximum, which was shifted to lower pH values with increasing buffer concentrations and increasing salting-out effect of the buffer anion according to the Hofmeister series. As this downshift correlates well with zeta potential values, a measurement of the latter allows the determination of the pH value for maximum DBC under a given set of conditions.