Enhanced Endotoxin Removal By Synergistic Effect of Positively Charged Depth Filtration and Polyethylene Glycol

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Sushma Ghimire, Philip Roppand 1 more

November 17, 2015

1 Min Read
Enhanced Endotoxin Removal By Synergistic Effect of Positively Charged Depth Filtration and Polyethylene Glycol

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Recombinant therapeutic proteins derived from E. coli require robust endotoxin removal, ideally performed using multiple process steps providing orthogonal and robust clearance. Typical recovery processes for E. coli expressed proteins consist of cell harvest by centrifugation, homogenization, collection of inclusion bodies by centrifugation, solubilization, refold, and clarification by depth filtration. Endotoxin clearance across the recovery operations is usually on the order of one log. Here, we demonstrate that endotoxin clearance can be enhanced to five logs by using 16% polyethylene glycol 4600 (PEG), a strong precipitating agent, in conjunction with positively charged depth filters. The highest dependence on endotoxin clearance was due to the amount of PEG utilized, while endotoxin clearance was less dependent on pore size and no dependence on charge type of depth filter. Thus, the endotoxin clearance observed across the depth filter appears to be most reliant on absorption followed by particulate removal. Previous work has demonstrated that PEG can increase the binding capacity of chromatography resins. Therefore, incorporating higher PEG concentration during depth filtration in the initial product recovery process can provide enhanced endotoxin clearance to the overall purification process.

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