Flow through anion exchange chromatography (AEX) has provided reliable process-and product-related impurities removal as well as viral clearance for monoclonal antibodies (mAbs). The application of AEX to molecules with more complex impurity profiles or non-platform characteristics such as a low pI can become challenging because viral clearance considerations often constrain the AEX step design space. Multimodal anion exchange chromatography (MMAEX) can address the limitations of the platform AEX step while still allowing a "platform-like" manufacturing process. This work presents a case study on polishing step development for a Fc-fusion protein with pI <
6.5, high surface hydrophobicity, and aggregate content of up to 35% in the harvested cell culture fluid. An integrated computational and experimental high throughput screening (HTS) workflow was implemented to rapidly identify the MMAEX resin Capto Adhere at pH 5.5 as a viable alternative to the conventional AEX flow through polishing step. This work presents the impact of pH, conductivity, load, and load impurity level/composition on MMAEX step performance. Step performance was measured based on yield, HMW clearance, residual HCP and DNA clearance, as well as viral clearance. HMW clearance varied based on the starting load HMW level (from 5 -35%) and the highest relative clearance was obtained at a moderate HMW load level. Residual HCP and DNA showed a strong dependance on load HMW level, highlighting the competitive adsorption behavior that can impact resin-protein interactions in complex mixtures. Both residual HCP and DNA were removed to below quantification at pH 5.5. Viral clearance of up to 4 logs of XMULV and MVM was demonstrated at pH 5.5 using process relevant feed streams. The workflow presented here demonstrates how the integration of in silico modeling and high throughput screening (HTS) can streamline process development and enable rapid polishing step optimization. These results also underscore the impact of feed impurity and impurity composition on MMAEX resin performance.