Separation apparatus and method
Protein separation can be provided based on a High Performance Countercurrent Membrane Purification (HPCMP), which can exploit highly selective diffusive transport across thin walls of a hollow fiber membrane for separation of proteins. Embodiments of an HPCMP system can separate mixtures of multiple proteins (e.g. separate BSA and Mb) or other biological material and provide high yields (e.g. achieving greater than 98% yield of both proteins with purification factors greater than 100-fold, etc.). Embodiments of a HPCMP system can be configured for high resolution separations in the preparation of biopharmaceuticals and natural protein products. Other embodiments can be utilized in other protein separation environments or biological material separation environments.
1 . A method of separating proteins comprising:
passing a feed to a High Performance Countercurrent Membrane Purification (HPCMP) module having at least one membrane within a vessel, the feed comprising a first protein and a second protein, the feed passed through the HPCMP module in countercurrent flow with a buffer fed to the vessel of the HPCMP module;
separating the first protein from the second protein via differences in diffusive flux across the at least one membrane of the HPCMP module;
outputting a product flow and a buffer outlet flow from the vessel of the HPCMP module, the product flow having a purity of the first protein that is greater than the purity of the first protein within the feed, the buffer outlet flow having a purity of the second protein that is greater than the purity of the second protein within the feed passed to the HPCMP module;
wherein the passing of the feed and the outputting of the product flow and the buffer outlet flow occur simultaneously during a continuous flow operation of the HPCMP module.
2 . The method of claim 1 , wherein the at least one membrane is a single use membrane.
3 . The method of claim 1 , wherein the at least one membrane is a hollow fiber membrane.
4 . The method of claim 1 , wherein the purity of the first protein within the product flow is at least fifty times higher than the purity of the first protein within the feed.
5 . The method of claim 1 , wherein the purity of the second protein within the buffer outlet flow is at least fifty times higher than the purity of the second protein within the feed.
6 . The method of claim 1 , wherein the purity of the first protein within the product flow is at least 100 times higher than the purity of the first protein within the feed and the purity of the second protein within the buffer outlet flow is at least 100 times higher than the purity of the second protein within the feed.
7 . The method of claim 1 , wherein the feed is a liquid flow and the buffer is a liquid, the product flow is a liquid and the buffer outlet flow is a liquid.
8 . The method of claim 1 , comprising:
outputting the feed from a bioreactor upstream of the HPCMP module;
passing the feed through at least one of ultrafiltration (UF) and dialysis upstream of the HPCMP module.
9 . The method of claim 8 , wherein the dialysis is countercurrent dialysis (CCD).
10 . The method of claim 8 , comprising:
passing the buffer through at least one of ultrafiltration (UF) and dialysis before the buffer is fed to the HPCMP module.
11 . The method of claim 10 , wherein the dialysis is countercurrent dialysis (CCD).
12 . The method of claim 1 , comprising:
controlling at least one of ionic strength of the feed and pH of the feed to increase selectivity.
13 . The method of claim 1 , comprising:
adjusting at least one of ionic strength of the buffer and pH of the buffer during the continuous operation of the HPCMP module to increase selectivity.
14 . The method of claim 1 in which the at least one membrane is pretreated to increase an effective pore size of the at least one membrane.
15 . The method of claim 1 , wherein the first protein is a monoclonal antibody, bovine serum albumin (BSA), myoglobin (Mb), or Immunoglobulin G (IgG); and
the second protein is a protein within a mixture of host cell proteins, BSA, Mb, or IgG.
16 . A method of separating biological material comprising:
passing a feed to a High Performance Countercurrent Membrane Purification (HPCMP) module having at least one membrane within a vessel, the feed comprising a first desired biological product and at least one second component, the feed passed through the HPCMP module in countercurrent flow with a buffer fed to the vessel of the HPCMP module;
separating the first desired biological product from the second component via differences in diffusive flux across the at least one membrane of the HPCMP module; and
outputting a product flow and a buffer outlet flow from the vessel of the HPCMP module, the product flow having a purity of the first desired biological product that is greater than the purity of the first desired biological product within the feed, the buffer outlet flow having a purity of the at least one second component that is greater than the purity of the at least one second component within the feed passed to the HPCMP module;
wherein the passing of the feed and the outputting of the product flow and the buffer outlet flow occur simultaneously during a continuous flow operation of the HPCMP module.
17 . The method of claim 16 , wherein the purity of the first desired biological product within the product flow is at least fifty times higher than the purity of the first desired biological product within the feed.
18 . The method of claim 16 , wherein the purity of the at least one second component within the buffer outlet flow is at least fifty times higher than the purity of the least one second component within the feed.
19 . The method of claim 16 , wherein the purity of the first desired biological product within the product flow is at least 100 times higher than the purity of the first desired biological product within the feed and the purity of the at least one second component within the buffer outlet flow is at least 100 times higher than the purity of the at least one second component within the feed.
20 . The method of claim 16 , wherein the first desired biological product is a pegylated protein, a glycoconjugate vaccine, a monoclonal antibody, bovine serum albumin (BSA), myoglobin (Mb), Immunoglobulin G (IgG) or an antibody drug conjugate; and
the at least one second component is unreacted polyethylene glycol, polysaccharide, or drug molecule or is a protein within a mixture of host cell proteins, BSA, Mb, or IgG.
21 . The method of claim 20 , wherein the first desired biological product is the monoclonal antibody, the bovine serum albumin (BSA), the myoglobin (Mb), or the Immunoglobulin G (IgG); and
the at least one second component is the protein within the mixture of host cell proteins, BSA, Mb, or IgG.