Most protein purification in biomanufacturing is currently performed using column chromatography. Although these systems can provide very high resolution separations, many chromatographic resins are quite expensive, scale-up can be challenging, and it is difficult to apply column chromatography in continuous downstream processes. Recent work in our lab has demonstrated the potential of High Performance Countercurrent Membrane Purification (HPCMP) for protein separations based on selective diffusive transport across low-cost hollow fiber membranes. The objective of the work to be presented in this talk was to explore opportunities to enhance HPCMP by: (1) tailoring the membrane pore size to target specific protein separations, and (2) exploiting the effects of buffer pH and ionic strength on protein partitioning / diffusion in porous membranes. Experiments were performed using commercially available hollow fiber membranes used for hemodialysis. These hollow fiber membranes were modified using bleach-hypochlorite solutions to increase the pore size for the purification of monoclonal antibodies, with human serum Immunoglobulin G (hIgG) used as a model. HPCMP separations were performed over a range of buffer pH and ionic strength, with the greatest selectivity obtained by operating at relatively low conductivity at pH ≈ 5, which is near the isoelectric point of albumin (used as a model impurity). A selectivity of 50-fold could be obtained, enabling one to purify the hIgG with 1000-fold removal of albumin at &gt;95% yield.  These results demonstrate the potential of using simple modification methods and buffer selection procedures to obtain high resolution proteins separations by HPCMP.

High performance countercurrent membrane purification (HPCMP)

ACS Spring 2022

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