Methods of protein fractionation using high performance tangential flow filtration
First Claim
1. A method for separating a protein of interest from a feedstream, comprising:
- (a) filtering said feedstream by a high performance tangential-flow filtration process that separates said molecular species of interest from said feedstream on the basis of pore size and charge of said protein of interest, while maintaining flux at a level ranging from about 5 to 100% of transition point flux in the pressure-dependent region of the flux versus TMP curve, wherein transmembrane pressure is held substantially constant along the membrane at a level no greater than the transmembrane pressure at the transition point of the filtration, whereby said protein of interest is selectively separated from said feedstream such that said protein of interest retains its biological activity;
(b) filtering said feedstream by an ultrafiltration process; and
, wherein said filtration is occurring above the transition point flux of said protein of interest;
wherein said molecular species has a molecular weight of between 1 and 1000 kDa.
5 Assignments
0 Petitions
Accused Products
Abstract
Processes and apparati are provided for separating molecules of interest from a mixture containing them which comprises subjecting the mixture to an improved method of high performance tangential flow filtration (HPTFF). The HPTFF of the invention was used to clarify, and process various feedstreams for the removal of a molecule of interest based on both the size and charge of the molecule of interest. According to a preferred embodiment, a transgenic milk feedstream is stabilized and particulate matter such as fat, casein miscelles and bacteria are removed. The method of HPTFF used in the current invention utilizes optimized process parameters that include temperature, trans-membrane pressure, molecular charge, molecular size, cross-flow velocity, and milk concentration. Cleaning and storage procedures were also developed to ensure long membrane life. An aseptic filtration step was also developed to remove any bacteria remaining in a clarified transgenic milk feedstream.
87 Citations
78 Claims
-
1. A method for separating a protein of interest from a feedstream, comprising:
-
(a) filtering said feedstream by a high performance tangential-flow filtration process that separates said molecular species of interest from said feedstream on the basis of pore size and charge of said protein of interest, while maintaining flux at a level ranging from about 5 to 100% of transition point flux in the pressure-dependent region of the flux versus TMP curve, wherein transmembrane pressure is held substantially constant along the membrane at a level no greater than the transmembrane pressure at the transition point of the filtration, whereby said protein of interest is selectively separated from said feedstream such that said protein of interest retains its biological activity;
(b) filtering said feedstream by an ultrafiltration process; and
,wherein said filtration is occurring above the transition point flux of said protein of interest;
wherein said molecular species has a molecular weight of between 1 and 1000 kDa. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 76, 77, 78)
-
-
75. A method for separating a protein of interest from a feedstream, comprising:
-
(a) filtering said feedstream by a high performance tangential-flow filtration process that separates said molecular species of interest from said feedstream on the basis of pore size and charge of said protein of interest, while maintaining flux at a level ranging from about 5 to 100% of transition point flux in the pressure-dependent region of the flux versus TMP curve, wherein transmembrane pressure is held substantially constant along the membrane at a level no greater than the transmembrane pressure at the transition point of the filtration, whereby said protein of interest is selectively separated from said feedstream such that said protein of interest retains its biological activity;
(b) filtering said feedstream by a microfiltration process; and
,(c) increasing transmembrane pressure and decreasing flux for the first half of the filtration;
(d) decreasing therafter increasing or maintaining flux as the filtration progresses. wherein said filtration is occurring above the transition point flux of said protein of interest;
wherein said molecular species has a molecular weight of between 1 and 1000 kDa.
-
Specification