Synergistic viral inactivation platform: Rapid enveloped virus inactivation at mildly acidic pH using low non-ionic detergent in Protein A eluates.

Low-pH viral inactivation (LPVI) is a widely used, validated unit operation for enveloped virus clearance in antibody manufacturing. However, acid exposure during Protein A (ProA) elution and subsequent low-pH holds can destabilize some antibodies and Fc-containing modalities, increasing aggregation risk and product loss-a challenge that is particularly acute for multi-specific antibody formats and other pH-labile Fc-containing modalities. Mild-pH ProA capture strategies protect such molecules, but the resulting higher-pH eluates often require re-acidification to meet conventional LPVI setpoints, reintroducing low-pH stress. Here we evaluate a dual-mechanism strategy in which mild acidity is combined with low concentrations of non-ionic detergent to accelerate inactivation without prolonged exposure to strongly acidic conditions. Using xenotropic murine leukemia virus (X-MuLV) spiked into ProA eluates, 0.10% N-methylglucamide (Mega-10) at pH 3.91 reduced infectivity to below the assay limit of detection within 2.5 min (log10 reduction factor [LRF] ≥ 5.40), whereas low pH alone or detergent alone produced substantially lower reductions over the same interval. Similarly, 0.15% Mega-10 at pH 4.05 achieved nondetectable infectivity within 10 min (LRF ≥ 5.81). Comparable time compression was observed with Tween 80 or Tween 20 at pH ~3.97, reducing time-to-nondetectable infectivity from 120 min (low pH alone) to 5 min. The operating window was tunable through paired adjustment of detergent concentration and pH, and feasibility was demonstrated at pH 4.56 with low detergent levels, offering a practical solution for pH-labile products and higher-pH eluates.