We have shown how protein self-association impacts the ion-exchange separation of modified forms and aggregates for apolipoprotein A-I(Milano). It is well known that reversible self-association of a protein can lead to chromatographic band broadening, peak splitting, merging, fronting, and tailing. To mitigate these effects, urea or an organic modifier can be added to the chromatography buffers to shift the equilibrium distribution of the target molecule to the dissociated form. A first generation process that did not utilize urea resulted in low yield and low purity as it was not possible to separate protein aggregates. A second generation process run in the presence of 6M urea resulted in high purity and high yield, but throughput was limited due to low resin binding capacity when the protein was completely denatured. A third generation process achieved high purity, high yield, and high throughput by shifting the urea concentration during the process to continually operate in the optimal window for maximum loading and selectivity. Key to these systematic process improvements was the rational understanding of the interplay of urea concentration and ion-exchange chromatographic behavior. Results from pilot and industrial scale operations are presented, demonstrating the suitability of the techniques described in this work for the large scale manufacture of recombinant therapeutic proteins.