Plasmid DNA (pDNA) is a critical starting material for the production of viral vectors, mRNA therapeutics, and recombinant proteins. The rapid growth of these modalities has increased demand for high-quality pDNA. Despite previous investigations of chaotropic additives and anion exchange chromatography for nucleic acid purification, significant plasmid losses remain a challenge in membrane-based processes. In this study, we developed and optimized a Mustang™ Q XT anion exchange membrane chromatography process to improve pDNA recovery directly from clarified lysate using a Design of Experiments (DOE) approach. Process parameters, including pH, conductivity, loading conditions, and buffer additives, were evaluated to identify factors affecting plasmid recovery and RNA removal. Arginine and urea were identified as significant process enhancers, increasing plasmid recovery from approximately 50% to as high as 93% while improving resolution between pDNA and RNA species. The additives also reduced the conductivity required for plasmid elution, improving recovery and chromatographic selectivity. Additional studies demonstrated that guanidine and histidine provided similar benefits. These improvements were achieved without additional chromatography or polishing steps, enabling direct processing of clarified lysate in a rapid, low-shear workflow. The findings provide a practical, scalable strategy for enhancing pDNA recovery using strong anion exchange membrane chromatography.
