Electrically driven elution in digital protein a membrane chromatography: An alternative to traditional low-pH...

Digital Protein A Membrane Chromatography (DMC^PrA) introduces an innovative antibody purification method that enables gentle elution using electrical fields rather than conventional low-pH buffers. By eliminating acidic conditions, DMC^PrA mitigates antibody degradation and aggregation associated with traditional Protein A chromatography. In this study, we investigated electrically driven elution in preparative Protein A membrane chromatography with the i3 DMC^PrA device (i3 Membrane GmbH, Radeberg, Germany). The effects of applied voltage, buffer composition, and salt type and concentration were systematically evaluated to clarify the underlying elution mechanisms. Effective elution occurred only with low-conductivity buffers and solutions, requiring specific salts at appropriate concentrations, for example 0.15 mM Na₂SO₄ or 1.0 mM NaCl, which yielded high antibody recovery. Applying voltage to induce elution, showed consistent pH shifts. When the electric field was oriented from anode to cathode, the pH increased, representing the optimal condition for voltage-driven elution; reversing polarity decreased the pH. The results support the hypothesis that the elution mechanism is primarily driven by electrochemical reactions at the electrodes, leading to water electrolysis. Hydroxide ions resulting from cathodic hydrogen evolution reactions (HER) increase pH, while protons from anodic oxygen evolution reactions (OER) likely induce conformational changes in Protein A and IgG, initiating elution. These reactions are strongly influenced by the applied voltage as well as the chemical composition and ionic properties of the buffers. Collectively, these parameters exert a significant impact on elution efficiency and reproducibility. This work demonstrates that electrically driven elution provides a promising alternative to low-pH based methods, enabling milder purification of antibodies through electrochemical modulation.