Prediction and validation of monoclonal antibodies separation in aqueous two-phase system using molecular dynamic...

In order to predict how mAbs partition in 20% ethylene oxide/80% propylene oxide (v/v) random copolymer (EO₂₀PO₈₀)/water aqueous two-phase system (ATPS), a molecular dynamic simulation model was developed using Gromacs and then validated by experiments. The ATPS was applied with seven kinds of salt, including buffer salt and strong dissociation salt that were commonly employed in the purification of protein. Na₂SO₄ was shown to have the best effects on lowering EO₂₀PO₈₀ content in the aqueous phase and enhancing recovery. The content of EO₂₀PO₈₀ in the sample solution was decreased to 0.62%±0.25% and the recovery of rituximab increased to 97.88%±0.95% by adding 300 mM Na₂SO₄ into back extraction ATPS. The viability determined by ELISA was 95.57% at the same time. A strategy for constructing a prediction model for the distribution of mAbs in ATPS was proposed in consideration of this finding. Partition of trastuzumab in ATPS was predicted by the model created using this method and the prediction result was further validated by experiments. The recovery of trastuzumab reached 95.63%±2.86% under the ideal extraction conditions suggested by the prediction model.