A capture step for pro-insulin expressed in E.Coli and an intermediate step for the purification of insulin following the capture step were developed. Resin screening and chromatography condition screening using an HTPD approach in a 96-well filter-plate format were performed.
The results indicated that the media of choice for the capture step was Capto™ MMC, a multi-modal cation-exchange resin. Elution conditions were further optimized in packed columns using the Design of Experiments (DoE) functionality in an ÄKTA™avant 25 chromatography system.
The optimal chromatographic conditions were identified and followed by a robustness study. The capture step was then scaled to pilot scale.
After enzymatic cleavage of the purified pro-insulin from the capture step the formed insulin was subjected to cation exchange chromatography. A similar approach for development of this step as for the capture step was followed. The results indicated that the best media for the intermediate step was Capto SP ImpRes. Robustness and scale up to pilot scale were performed.