In a paper titled „Electrically driven elution in digital Protein A membrane chromatography: An alternative to traditional low-pH elution“, the research group led by Prof. Dr. Christian Frech at the Institute of Biochemistry at Mannheim University of Applied Sciences investigates how electric fields can be used for the elution of monoclonal antibodies in i3 Digital Membrane Chromatography (DMC).1
An alternative to conventional low-pH elution
Protein A chromatography is a key step in the purification of monoclonal antibodies and antibody variants. In conventional processes, elution typically occurs using acidic buffers in the pH range of 3 to 4. These conditions can stress sensitive antibodies and lead to aggregation or other molecular changes.
The published study investigates an alternative approach: electrically driven elution using Protein A membranes. In this method, antibody elution is not triggered by a conventional low-pH elution buffer, but rather by applying an electric field. The results show that efficient voltage-induced elution is possible under defined low-conductivity conditions. In the experiments conducted, antibody recoveries of over 95% were achieved. The subsequent acidic control elution revealed only small residual amounts of antibody, indicating that the vast majority had already been released during the electrical elution.
Initial Insights into the DMC-Mechanism
The study also provides a mechanistic explanation for the observed pH changes during electrically driven elution. The authors attribute these pH shifts to electrochemical reactions at the electrodes, specifically the oxygen evolution reaction at the anode and the hydrogen evolution reaction at the cathode. These reactions result in the local formation of H₃O⁺ and OH⁻ ions.
Although an alkaline bulk pH was frequently measured at the system’s outlet, it is assumed that a local drop in pH can occur within the Protein A membrane. This local effect could dissociate the Protein A/antibody interaction and thereby enable elution. Thus, the study provides a plausible explanation for how Protein A elution can be achieved without a conventional acidic elution buffer.
Further control experiments support this interpretation. The observed pH profiles also occurred in the absence of antibody loading and with ligand-free membranes. This demonstrates that the pH shifts are not specifically caused by Protein A or the antibody, but are attributable to general electrochemical effects within the system.
The experimental data and detailed results are available in the original publication.
Conductivity and Electrolyte Composition as Key Process Parameters
Another focus of the publication is the influence of buffer conductivity and electrolyte composition. The study shows that efficient elution does not depend solely on the applied voltage, but rather significantly on the properties of the buffer used. While undiluted PBS did not allow for sufficient voltage-driven elution, high elution efficiencies were achieved under low-conductivity conditions. The decisive factor here is not a specific ion such as chloride, but rather the conductivity of the entire electrolyte system.
Conductivity as a Key Efficiency Factor
Another focus of the publication is the influence of buffer conductivity and electrolyte composition. The study shows that efficient elution does not depend solely on the applied voltage, but rather significantly on the properties of the buffer used. While undiluted PBS did not allow for sufficient voltage-driven elution, high elution efficiencies were achieved under low-conductivity conditions. The decisive factor here is not a specific ion such as chloride, but rather the conductivity of the entire electrolyte system.
Scalability of Voltage-Driven Elution
The study also showed that voltage-driven elution can be scaled up to a larger device format with a 1 mL membrane volume and a binding capacity of up to 50 mg hIgG. By adjusting the voltage and application duration, an elution efficiency of over 95% was achieved. Furthermore, targeted polarity reversal enabled the achievement of nearly neutral eluate conditions, yielding 8.0 mL of eluate at pH 7.37. Thus, the results demonstrate that electrically driven Protein A elution is reproducible, mechanistically explainable, and scalable under suitable process conditions.
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