Affinity chromatography device comprising a fibrillated heat-treated polymer membrane and concentrator comprising the affinity chromatography device

Abstract

Disclosed are affinity chromatography devices comprising a fibrillated heat treated polymeric membrane containing inorganic particles having a spherical shape and a particle size distribution with D90 / D10 less than or equal to 3. A blend or composition of spherical inorganic particles having a nominal particle size of about 5 microns to about 20 microns can be used. Further, the devices have a hydraulic permeability of about 200 (X 10 ‑12 cm 2 ) to about 700 (X 10 ‑12 cm 2 ). The affinity chromatography devices have a dynamic binding capacity (DBC) at 10% breakthrough of greater than 35 mg / ml at a residence time of 20 seconds. Additionally, the affinity chromatography devices have a cycle durability of at least 100 cycles at an operating pressure of no more than 0.3 MPa. Also disclosed are collectors comprising a plurality of affinity chromatography devices and a plurality of collectors in parallel configuration.

Description

Technical Field

[0001] This disclosure generally relates to affinity chromatography, and more specifically to an affinity chromatography apparatus comprising a fibrillated heat-treated polymer membrane containing spherical particles having a particle size distribution of D90 / D10 less than or equal to 3, and capable of separating target molecules from aqueous mixtures. Manifolds comprising multiple affinity chromatography apparatuses and multiple manifolds configured in parallel are also disclosed. Background Technology

[0002] Chromatographic methods are commonly used to separate and / or purify molecules of interest, such as proteins, nucleic acids, and polysaccharides, from mixtures. Affinity chromatography specifically involves passing a mixture through a matrix containing a specific ligand (i.e., a specific binding partner) that binds to the molecule of interest. Once exposed to the ligand, the molecule of interest binds to the matrix and is thus retained from the mixture. Affinity chromatography offers several advantages over other types of chromatography. For example, it provides a purification method that can separate the target protein from a mixture of other biomolecules in a single step with high yields.

[0003] Despite the many advantages of current affinity chromatography devices, there is a need in the art for a chromatography device that can be used for shorter residence times than conventional devices, while providing the same or better binding capacity as current products, and that can be reused. Summary of the Invention

[0004] In one aspect ("Aspect 1"), an affinity chromatography apparatus includes a fluid inlet, a fluid outlet fluidly connected to the fluid inlet, a protofibrotic heat-treated polymer membrane (containing inorganic particles having a spherical shape and a nominal particle size of about 5 micrometers to about 20 micrometers) located between the fluid inlet and the fluid outlet, a shell surrounding the fluid inlet, the fluid outlet, and the protofibrotic heat-treated polymer membrane, wherein the particle size distribution has a D90 / D10 of less than or equal to 3, and wherein at least one of the protofibrotic polymer membrane and the inorganic particles is covalently bonded to an affinity ligand, the affinity ligand reversibly binding to a target molecule.

[0005] According to another aspect of aspect 1 ("Aspect 2"), wherein the target molecule is a protein, antibody, viral vector, or combination thereof.

[0006] According to another aspect of aspect 1 or 2 ("Aspect 3"), approximately 200 (X 10) -12 cm 2 ) to approximately 700 (X 10) -12 cm 2Hydraulic penetration rate.

[0007] According to another aspect of any of aspects 1 to 3 ("Aspect 4"), the inorganic particles having a spherical shape are selected from silica, zeolite, hydroxyapatite, metal oxides and combinations thereof.

[0008] According to another aspect of any of aspects 1 to 4 ("Aspect 5"), the fibrillated polymer membrane includes expanded polytetrafluoroethylene membrane, expanded modified polytetrafluoroethylene membrane, expanded polytetrafluoroethylene copolymer membrane or expanded polyethylene membrane.

[0009] According to another aspect of any of aspects 1 to 5 ("Aspect 6"), the protofibrillated heat-treated polymer film is an expanded polytetrafluoroethylene film.

[0010] According to another aspect of any of aspects 1 to 6 ("Aspect 7"), the affinity ligand is selected from protein A, protein G, protein L, human Fc receptor protein, antibody, polysaccharide and combination thereof.

[0011] According to another aspect of any of aspects 1 to 7 ("Aspect 8"), the inorganic particles include at least a first inorganic particle having a spherical shape and a first nominal particle size and a second inorganic particle having a spherical shape and a second nominal particle size, the first nominal particle size and the second nominal particle size being different from each other.

[0012] According to another aspect of any of aspects 1 to 8 ("Aspect 9"), the nominal particle size is selected from about 5 micrometers, about 10 micrometers, about 15 micrometers, about 20 micrometers, and combinations thereof.

[0013] According to another aspect of any of aspects 1 to 9 ("Aspect 10"), the inorganic particles having a spherical shape include a blend of 10-micrometer spherical particles and 20-micrometer spherical particles, and the blend is 10:90 to 90:10.

[0014] According to another aspect of any of aspects 1 to 9 ("Aspect 11"), the inorganic particles having a spherical shape include a blend of spherical particles of 5 micrometers and spherical particles of 10 micrometers, and the blend is 10:90 to 90:10.

[0015] According to another aspect of any of aspects 1 to 9 ("Aspect 12"), the inorganic particles having a spherical shape include a blend of spherical particles of 5 micrometers and spherical particles of 20 micrometers, and the blend is 10:90 to 90:10.

[0016] According to another aspect of any of aspects 1 to 12 ("Aspect 13"), it includes a dynamic binding capacity (DBC) of at least 35 mg / ml at a residence time of 20 seconds.

[0017] According to another aspect of any of aspects 1 to 13 ("Aspect 14"), it includes cyclic durability of at least 100 cycles at an operating pressure of less than 0.3 MPa.

[0018] According to another aspect of any of aspects 1 to 14 ("Aspect 15"), the protofibrotic heat-treated polymer film has a wound structure.

[0019] According to another aspect of any of aspects 1 to 15 ("Aspect 16"), the fibrillated polymer membrane has a stacked structure.

[0020] According to another aspect of any of aspects 1 to 16 ("Aspect 17"), the fibrillated polymer membrane has a wound structure, a stacked structure, and a combination thereof.

[0021] According to another aspect of any of aspects 1 to 17 ("Aspect 18"), wherein the inner intermediate material surrounds the outer surface of the core, and wherein the protofibrous polymer membrane surrounds the inner intermediate material.

[0022] According to another aspect of any of aspects 1 to 18 ("Aspect 19"), it includes an outer intermediate material surrounding the original fibrous polymer membrane.

[0023] According to another aspect of any of aspects 1 to 19 ("Aspect 20"), the inner intermediate material and the outer intermediate material are selected from porous fluoropolymer membranes, porous non-fluoropolymer membranes, porous nonwoven materials and porous woven materials.

[0024] According to another aspect of any of aspects 1 to 20 ("Aspect 21"), at least one of the inner layer intermediate material and the outer layer intermediate material is a polypropylene nonwoven material.

[0025] In one aspect ("Aspect 22"), an aggregator comprises at least two affinity chromatography devices arranged in parallel configurations as described in any one of Aspects 1 to 21.

[0026] According to another aspect of aspect 23 (“Aspect 23”), the collector is enclosed in a housing.

[0027] According to another aspect ("Aspect 24"), an article of any of Aspects 1 to 21 is used to separate a target substance from a biological sample.

[0028] In one aspect ("Aspect 25"), an apparatus includes a first collector and a second collector constructed in parallel, wherein each of the first collector and the second collector includes at least a plurality of affinity chromatography devices as described in any of aspects 1 to 21.

[0029] According to another aspect of aspect 25 ("Aspect 26"), the first collector and the second collector are enclosed in a housing.

[0030] According to one aspect ("Aspect 27"), an article comprising a core located at a center, a protofibrotic heat-treated polymer film (containing spherical inorganic particles having a spherical shape and a nominal particle size of about 5 micrometers to about 20 micrometers) wound around the core, a shell surrounding the core and the protofibrotic heat-treated polymer film, a first end cap disposed at a first end of the shell, and a second end cap disposed at a second end of the shell, the particle size distribution having a D90 / D10 of less than 3, and wherein at least one of the protofibrotic heat-treated polymer film and the spherical inorganic particles is covalently bonded to an affinity ligand, the affinity ligand reversibly binding to a target molecule.

[0031] According to another aspect of aspect 27 ("Aspect 28"), wherein the target molecule is a protein, antibody, viral vector, or combination thereof.

[0032] According to another aspect of aspect 27 or 28 ("Aspect 29"), it includes approximately 200 (X 10) 12 cm 2 ) to approximately 700 (X10) 12 cm 2 Hydraulic penetration rate.

[0033] According to another aspect of any of aspects 27 to 29 ("Aspect 30"), wherein the inorganic particles having a spherical shape are selected from silica, zeolite, hydroxyapatite, metal oxides and combinations thereof.

[0034] According to another aspect of any of aspects 27 to 30 ("Aspect 31"), the fibrillated polymer membrane comprises at least one of expanded polytetrafluoroethylene membrane, expanded modified polytetrafluoroethylene membrane, expanded polytetrafluoroethylene copolymer membrane or expanded polyethylene membrane.

[0035] According to another aspect of any of aspects 27 to 31 ("Aspect 32"), the protofibrillated heat-treated polymer membrane is a protofibrillated heat-treated expanded polytetrafluoroethylene membrane.

[0036] According to another aspect of any of aspects 27 to 32 ("Aspect 33"), the affinity ligand is selected from protein A, protein G, protein L, human Fc receptor protein, antibody, polysaccharide and combination thereof.

[0037] According to another aspect of any of aspects 27 to 33 ("Aspect 34"), the inorganic particles include at least a first inorganic particle having a spherical shape and a first nominal particle size and a second inorganic particle having a spherical shape and a second nominal particle size, and wherein the first nominal particle size and the second nominal particle size are different from each other.

[0038] According to another aspect of any of aspects 27 to 34 ("Aspect 35"), the nominal particle size is selected from about 5 micrometers, about 10 micrometers, about 15 micrometers, about 20 micrometers, and combinations thereof.

[0039] According to another aspect of any of aspects 27 to 35 ("Aspect 36"), the inorganic particles having a spherical shape comprise a blend of 10-micrometer spherical particles and 20-micrometer spherical particles, wherein the blend is 10:90 to 90:10.

[0040] According to another aspect of any of aspects 27 to 35 ("Aspect 37"), the inorganic particles having a spherical shape include a blend of 5-micrometer spherical particles and 10-micrometer spherical particles, and the blend is 10:90 to 90:10.

[0041] According to another aspect of any of aspects 27 to 35 ("Aspect 38"), the inorganic particles having a spherical shape comprise a blend of 5-micrometer spherical particles and 20-micrometer spherical particles, and the blend is 10:90 to 90:10.

[0042] According to another aspect of any of aspects 27 to 38 ("Aspect 39"), it includes a dynamic binding capacity (DBC) of at least 35 mg / ml at a residence time of 20 seconds.

[0043] According to another aspect of any of aspects 27 to 39 ("Aspect 40"), it includes cycle durability of at least 100 cycles and operating pressure of less than 0.3 MPa.

[0044] According to another aspect of any of aspects 27 to 40 ("Aspect 41"), the inner intermediate material surrounds the outer surface of the core, and the protofibrotic heat-treated polymer film surrounds the inner intermediate material.

[0045] According to another aspect of any of aspects 27 to 41 ("Aspect 42"), it includes an outer intermediate material surrounding the original fibrous heat-treated polymer film.

[0046] According to another aspect of any of aspects 27 to 42 ("Aspect 43"), the inner intermediate material and the outer intermediate material are selected from porous fluoropolymer membranes, porous non-fluoropolymer membranes, porous nonwoven materials and porous woven materials.

[0047] According to another aspect of any of aspects 27 to 43 ("Aspect 44"), at least one of the inner layer intermediate material and the outer layer intermediate material is a polypropylene nonwoven material.

[0048] In one aspect ("Aspect 45"), an aggregator includes at least two affinity chromatography devices arranged in parallel configurations, such as those in any of Aspects 27 to 44.

[0049] According to another aspect of aspect 45 (“Aspect 46”), the collector is enclosed in a housing.

[0050] According to one aspect ("Aspect 47"), an apparatus includes a first collector and a second collector constructed in parallel, wherein each of the first collector and the second collector includes at least two affinity chromatography devices as described in any one of aspects 27 to 44.

[0051] According to another aspect of aspect 47 ("Aspect 48"), the first collector and the second collector are enclosed in a housing.

[0052] According to another aspect ("Aspect 49"), an article of any of aspects 27 to 44 is used to separate target molecules from a fluid stream.

[0053] According to one aspect (Aspect 50), an affinity chromatography apparatus includes a housing, an inlet for fluid inflow into the housing member, first and second fluid distributors (the first fluid distributor and the second fluid distributor are located at opposite ends of the housing), an outlet for fluid outflow from the housing, and a stacked membrane assembly disposed within the housing between the fluid inlet and the fluid outlet, the stacked membrane assembly comprising: two or more layers of a stacked protofibrotic heat-treated polymer membrane, the protofibrotic heat-treated polymer membrane containing inorganic particles having a spherical shape and a nominal particle size of about 5 micrometers to about 20 micrometers, wherein the particle size distribution has a D90 / D10 of less than 3, and at least one of the protofibrotic heat-treated polymer membrane and the spherical inorganic particles is covalently bonded to an affinity ligand, the affinity ligand reversibly binding to a target molecule.

[0054] According to another aspect of aspect 50 ("Aspect 51"), wherein the target molecule is a protein, antibody, viral vector, or combination thereof.

[0055] According to another aspect of aspect 50 or 51 ("Aspect 52"), it includes first and second fluid distributors, the first fluid distributor and the second fluid distributor being located at opposite ends of the housing.

[0056] According to another aspect of any of aspects 50 to 52 ("Aspect 53"), it includes approximately 200 (x 10) -12 cm 2 ) to approximately 700 (X 10) -12 cm 2 Hydraulic penetration rate.

[0057] According to another aspect of any of aspects 50 to 53 ("Aspect 54"), wherein the inorganic particles having a spherical shape are selected from silica, zeolite, hydroxyapatite, metal oxides and combinations thereof.

[0058] According to another aspect of any of aspects 50 to 54 ("Aspect 55"), the fibrillated polymer membrane includes expanded polytetrafluoroethylene membrane, expanded modified polytetrafluoroethylene membrane, expanded polytetrafluoroethylene copolymer membrane or expanded polyethylene membrane.

[0059] According to another aspect of any of aspects 50 to 55 ("Aspect 56"), the protofibrillated heat-treated polymer membrane is a protofibrillated heat-treated expanded polytetrafluoroethylene membrane.

[0060] According to another aspect of any of aspects 50 to 56 ("Aspect 57"), the affinity ligand is selected from protein A, protein G, protein L, human Fc receptor protein, antibody, polysaccharide and combination thereof.

[0061] According to another aspect of any of aspects 50 to 57 ("Aspect 58"), the inorganic particles include at least a first inorganic particle having a spherical shape and a first nominal particle size and a second inorganic particle having a spherical shape and a second nominal particle size, wherein the first nominal particle size and the second nominal particle size are different from each other.

[0062] According to another aspect of any of aspects 50 to 58 ("Aspect 59"), the nominal particle size is selected from about 5 micrometers, about 10 micrometers, about 15 micrometers, about 20 micrometers, and combinations thereof.

[0063] According to another aspect of any of aspects 50 to 59 ("Aspect 60"), the inorganic particles having a spherical shape comprise a blend of 10-micrometer spherical particles and 20-micrometer spherical particles, and the blend is 90:10 to 10:90.

[0064] According to another aspect of any of aspects 50 to 59 ("Aspect 61"), the inorganic particles having a spherical shape include a blend of 5-micrometer spherical particles and 10-micrometer spherical particles, and the blend is 10:90 to 90:10.

[0065] According to another aspect of any of aspects 50 to 59 ("Aspect 62"), the inorganic particles having a spherical shape comprise a blend of spherical particles of 5 micrometers and spherical particles of 20 micrometers, and wherein the blend is 10:90 to 90:10.

[0066] According to another aspect of any of aspects 50 to 62 ("Aspect 63"), it includes a dynamic binding capacity (DBC) of at least 35 mg / ml at a residence time of 20 seconds.

[0067] According to another aspect of any of aspects 50 to 63 ("Aspect 64"), wherein the cycle durability is at least 100 cycles and the operating pressure is less than 0.3 MPa.

[0068] According to another aspect of any of aspects 50 to 64 ("Aspect 65"), wherein at least one upper intermediate material is located on a first side of the stacked membrane assembly and a lower intermediate material is located on a second side of the stacked membrane assembly, the second side being opposite to the first side.

[0069] According to another aspect of aspect 65 ("Aspect 66"), the first intermediate material and the second intermediate material are selected from porous fluoropolymer membranes, porous non-fluoropolymer membranes, porous nonwoven materials and porous woven materials.

[0070] According to another aspect of aspect 66 ("Aspect 67"), at least one of the first intermediate material and the second intermediate material is a polypropylene nonwoven material.

[0071] In one aspect ("Aspect 68"), an aggregator includes at least two affinity chromatography devices arranged in parallel configurations, such as those in any of Aspects 50 to 67.

[0072] According to another aspect of aspect 68 ("Aspect 69"), the collector is enclosed in a housing.

[0073] In one aspect ("Aspect 70"), an apparatus includes a first collector and a second collector constructed in parallel, wherein each of the first collector and the second collector includes at least two affinity chromatography devices as described in any one of aspects 50 to 69.

[0074] According to another aspect of aspect 70 (“Aspect 71”), the first collector and the second collector are enclosed in a housing.

[0075] In another aspect ("Aspect 72"), an article of manufacture includes a diagnostic device comprising a protofibrotic polymer membrane located between a fluid inlet and a fluid outlet and containing inorganic particles having a spherical shape and a nominal particle size of about 5 micrometers to about 20 micrometers, wherein the particle size distribution has a D90 / D10 of less than or equal to 3, and wherein at least one of the protofibrotic polymer membrane and the inorganic particles is covalently bonded to a ligand that reversibly binds to a target molecule in a biological sample.

[0076] According to another aspect of aspect 72 ("Aspect 73"), it includes a fluid inlet and a fluid outlet fluidly connected to the fluid inlet.

[0077] According to another aspect of aspect 72 or 73 ("Aspect 74"), it includes a housing component surrounding the fluid inlet, the fluid outlet, and the original fibrous polymer membrane.

[0078] Brief description of the attached figures

[0079] The accompanying drawings, which are incorporated in and form part of this specification, are used to aid in a further understanding of this disclosure and illustrate embodiments thereof, and together with the specification, serve to explain the principles of this disclosure.

[0080] Figure 1 This is an exploded view of a chromatographic apparatus according to at least one embodiment, the apparatus including a wound membrane assembly comprising a protofibrous heat-treated polymer membrane containing spherical inorganic polymer particles.

[0081] Figure 2 This is a cross-sectional view of a chromatographic apparatus according to at least one embodiment, depicting the outer flow channel and the inner flow channel according to at least one embodiment;

[0082] Figure 3 This is a schematic cross-sectional view of another helical winding, vertical (normal) flow chromatography apparatus according to at least one embodiment.

[0083] Figure 4 This is an exploded view of a chromatographic apparatus containing a stacked membrane assembly according to at least one embodiment, the assembly including a fibrillated heat-treated polymer membrane containing spherical inorganic particles.

[0084] Figure 5 It is a diagram illustrating the relationship between the dynamic binding capacity (DBC) and liquid permeability of the affinity chromatography apparatus described in Example 1 according to at least one embodiment;

[0085] Figure 6 It is a diagram illustrating the relationship between the dynamic binding capacity (DBC) and liquid permeability of the affinity chromatography apparatus described in Example 2 according to at least one embodiment;

[0086] Figure 7 It is a schematic diagram of the front view of an aggregator containing two parallel chromatographic devices.

[0087] Figure 8 yes Figure 7 A schematic diagram of the top view of the collector;

[0088] Figure 9 It is a schematic perspective view of an collector containing four parallel chromatographic devices;

[0089] Figure 10 yes Figure 9 A schematic diagram of the top view of the collector;

[0090] Figure 11 It is a schematic diagram of the front view of two collectors arranged in parallel structures;

[0091] Figure 12 yes Figure 11 A schematic diagram of the top view of the two collectors; Detailed Implementation

[0092] Those skilled in the art will readily understand that various aspects of this disclosure can be implemented by any number of methods and apparatuses configured to perform the desired functions. It should also be noted that the accompanying drawings referenced herein are not necessarily drawn to scale but may be enlarged to illustrate various aspects of this disclosure, and in this regard, the drawings should not be considered limiting. It should be understood that, as used herein, the term "on" means one element (such as a polymer membrane) directly on another element, or there may be intermediate elements present. It should be understood that the terms "spherical particles," "spherical inorganic particles," and "inorganic particles having a spherical shape" are used interchangeably herein. Furthermore, the term "spiral-wound membrane assembly" refers to a combination of a separate fibrillated heat-treated polymer membrane and a fibrillated heat-treated polymer membrane with an intermediate material. Furthermore, the term "stacked membrane assembly" refers to a combination of a separate fibrillated heat-treated polymer membrane and a fibrillated heat-treated polymer membrane with one or more intermediate materials. The terms "fibrillated heat-treated polymer membrane" and "heat-treated fibrillated polymer membrane" are used interchangeably herein. Furthermore, the terms "affinity chromatography apparatus" and "chromatographic apparatus" are used interchangeably herein.

[0093] This disclosure relates to an affinity chromatography apparatus that separates target molecules from an aqueous mixture containing the target molecules. Target molecules include, but are not limited to, proteins, antibodies, viral vectors, and combinations thereof. The affinity chromatography apparatus includes a protofibrotic heat-treated polymer membrane containing inorganic particles having a spherical shape and a particle size distribution with a D90 / D10 ratio less than or equal to 3. In some embodiments, blends or compositions of spherical inorganic particles of various sizes are used. The nominal particle size of the spherical inorganic particles is from about 5 micrometers to about 20 micrometers. Affinity ligands may be bonded to the spherical inorganic particles and / or to the protofibrotic heat-treated polymer membrane. The heat treatment of the polymer membrane can result in a significant and beneficial increase in the liquid permeability of the apparatus, thereby achieving a desired shorter residence time while maintaining acceptable apparatus pressure. While the heat treatment provides the benefit of higher permeability, it causes a proportionally small decrease in the apparatus binding capacity (DBC). At a residence time of 20 seconds, the dynamic binding capacity of the chromatography apparatus at 10% breakthrough is greater than 35 mg / ml. Furthermore, this affinity chromatography device exhibits cycle durability of at least 100 cycles at an operating pressure not exceeding 0.3 MPa. It should be understood that the term "about" as used herein signifies + / - 10% of the specified unit of measurement.

[0094] look Figure 1 and Figure 2The document describes a wound chromatography apparatus 100. In forming the wound chromatography apparatus 100, at least one layer of a fibrillated, heat-treated polymer membrane containing spherical inorganic particles is wound around a cylindrical core 150. As used herein, fibrillation refers to the inclusion of fibrils in a polymer membrane, for example, a membrane with a microstructure characterized by nodes interconnected by fibrils, where voids are spaces between the nodes and fibrils. As used herein, "heat-treated" refers to the final product of a heat-treatment step in which the fibrillated polymer membrane is subjected to a temperature above its melting point for a specified time period. In some embodiments, at least one inner intermediate material 205 may be circumferentially positioned around (e.g., wind around) the core 150 to achieve a desired width or predetermined amount. The protofibrotic heat-treated polymer film 210, containing spherical inorganic particles, is then wound around the core 150 onto the inner intermediate material 205 to achieve a desired width or predetermined amount, and at least one outer intermediate material 225 is wrapped (e.g., wound) around the protofibrotic heat-treated polymer film 210 to achieve a desired width or predetermined amount. In this document, the combination of the inner intermediate material 205, the protofibrotic heat-treated polymer film 210, and the outer intermediate material 225 will be referred to as a "wound film assembly." In some embodiments, a "wound film assembly" may include a protofibrotic heat-treated polymer film and one or more inner intermediate materials, a protofibrotic heat-treated polymer film and one or more outer intermediate materials, and any combination of polymer and intermediate materials wound around the core. The cylindrical core 150 may have a hollow or solid interior. In either case, the core 150 includes a solid outer wall, such that the aqueous mixture flowing through the wound chromatography apparatus 100 flows within an inner channel formed by one or more inner intermediate layers of material, which will be discussed in detail below. Using a hollow core 150 reduces the amount of material used to form the core 150, lightens the weight of the wound chromatography apparatus 100, and lowers manufacturing costs.

[0095] The membrane module 110 and the central core 150 may be located within the housing member 50. In some embodiments, the housing member 50 is cylindrical. Figure 1 and Figure 2In the embodiments described herein, the outer intermediate material 225 forms the outer flow channel 130, and the inner intermediate material 205 forms the inner flow channel 140. It should be understood that the intermediate materials 205 and 225 in the embodiments described herein may be different or the same. Furthermore, one or more intermediate materials may be used to form one or both of the outer flow channel 130 and the inner flow channel 140. In use, an aqueous mixture flows into the inlet 80 located within the inlet end cap 60, whereby the mixture flows through the distributor cap 65 and is directed to the outer flow channel formed by the outer intermediate material 225. The distributor cap 65 directs the aqueous mixture 90 degrees from the feed direction to the outer flow channel 130 (i.e., one or more layers of intermediate material 225). This redirection promotes a more uniform flow of the aqueous mixture into the outer flow channel 130. The outer intermediate material 225 forming the outer flow channel 130 is located between the housing member 50 and the wound polymer film 210. The distributor cap 65 may be polyolefin or coated with polyolefin. It should be understood that the aqueous mixture flows along the outer channel gap 165 and connects to the outer flow channel 130.

[0096] The aqueous mixture flows through the outer channel 130 (i.e., one or more outer intermediate materials 225) in a vertical direction (e.g., vertical flow) across the wound polymer membrane 210. As the aqueous mixture flows vertically through the wound polymer membrane 210 from the outer channel 130 (i.e., the outer intermediate material 225), the affinity ligands reversibly bind to the target protein, thereby effectively removing it from the aqueous mixture. The aqueous mixture then enters the inner channel 140 (i.e., one or more inner intermediate materials 205) located between the solid outer wall of the central core 150 and the wound polymer membrane 210.

[0097] The aqueous mixture is then redirected at the bottom of the inner flow channel 140 by the outlet end cap 75. The aqueous mixture then flows out of the wound chromatography apparatus 100 through the outlet 85 located within the outlet end cap 75. It should be understood that the diameter and / or height of the central core 150 (and / or the width and / or height of the protofibrotic heat-treated polymer membrane and / or intermediate material) can be adjusted to achieve greater capacity without negatively impacting the performance of the apparatus. Furthermore, target proteins can be removed from affinity ligands, for example, by passing a liquid with a lower pH through the chromatography apparatus, as is known to those skilled in the art.

[0098] There are no particular limitations on intermediate materials 205 and 225, provided that the aqueous mixture can flow through them. Some non-limiting examples of suitable intermediate materials include, but are not limited to, porous fluoropolymer membranes or porous non-fluoropolymer membranes (e.g., porous polypropylene or other porous polyolefin membranes), porous nonwoven materials, or porous woven materials. In some embodiments, the wound membrane assembly includes an integrated inlet end cap 60 at one end of the core 150 and an integrated outlet end cap 75 at the other end of the core 150 to form an integrated, reusable chromatography device.

[0099] There is no particular limitation on the total number of fibrillated polymer membrane layers present in a wound membrane module, and it depends on the desired end use and / or the desired mass transit flow within the membrane module. Wound membrane modules may include 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 (or more) polymer membrane layers. It should be understood that hundreds or even thousands of polymer membrane layers may be present in a stacked membrane module. Furthermore, the fibrillated polymer membrane present in the wound membrane module may have a single-layer thickness of about 1 micrometer to about 10,000 micrometers, about 100 micrometers to about 5,000 micrometers, about 500 micrometers to about 3,000 micrometers, or about 650 micrometers to about 1,000 micrometers. As used herein, the term "thickness" refers to the direction of the fibrillated polymer membrane perpendicular to its length-area.

[0100] Figure 3 The function of the chromatographic apparatus 300 described in the text is the same as... Figure 2 The spiral wound chromatography apparatus 100 described herein is substantially similar. For example, an aqueous mixture is introduced into the chromatography apparatus 300 through an inlet 80 disposed within an inlet end cap 60 in the direction of arrow 62. The aqueous mixture is guided from the feed direction by a dispenser cap 65. For ease of description, the aqueous mixture may flow toward the outer channel 130 (which may be formed of an outer intermediate material 225) in the direction described by arrow 55. The aqueous mixture flows along the outer channel gap 165 and connects to the inner channel 140, in which it flows in the direction of arrow 30. The aqueous mixture flows through the spiral wound polymer membrane 210 in a direction perpendicular to arrow 70 (e.g., vertical flow) from the outer channel 130 to the inner channel 140. As the aqueous mixture passes through the spiral wound polymer membrane 210, affinity ligands reversibly bind to the target molecule. It should be understood that the inner channel 140 may be formed of one or more inner intermediate materials 205.

[0101] The aqueous mixture, free of the target molecule, flows along the inner flow channel 140 in the direction described by arrow 40. At the bottom of the inner flow channel 140, the aqueous mixture is redirected, flowing towards the central portion of the chromatography apparatus 300 as described by arrow 52. The aqueous mixture, free of the target molecule, flows out of the chromatography apparatus 300 through the outlet 85 within the outlet end cap 75 in the direction of arrow 45.

[0102] In other implementations, such as Figure 4 As described, the chromatographic apparatus 200 includes a fibrillated heat-treated polymer membrane configured such that individual fibrillated heat-treated polymer membrane discs 240 are stacked together to form a stacked membrane assembly 220. The stacked fibrillated heat-treated polymer membrane discs 240 can be formed by simply stacking them together. Alternatively, the fibrillated heat-treated polymer membrane discs 240 can be stacked and then laminated together using heat and / or pressure or other conventional methods. It should be understood that, for ease of interpretation, the stacked membrane assembly described herein is relative to the fibrillated heat-treated polymer membrane disc 240. Fibrillated polymer membranes formed in one or more geometries and / or one or more non-geometric shapes are considered to fall within the scope of this disclosure.

[0103] The chromatography apparatus 200 includes at least one upper intermediate material 260 disposed on top of the stacked membrane assembly 220 and at least one lower intermediate material 280 disposed below the stacked membrane assembly 220. The upper and lower intermediate materials 260 and 280 may be the same or different. Similar to the wound membrane assembly discussed above, there are no particular limitations on the intermediate materials 260 and 280 used to form the stacked membrane assembly 220, as long as an aqueous mixture can flow through it. Non-limiting examples of suitable intermediate materials include, but are not limited to, porous fluoropolymer membranes or porous non-fluoropolymer membranes (e.g., porous polypropylene or other porous polyolefin membranes), porous nonwoven materials, or porous woven materials.

[0104] The stacked membrane assembly 220 can be housed within a housing 250, which has an inlet cap 265 and an outlet cap 275 at opposite ends. In some embodiments, the housing 250 is cylindrical, although any geometry capable of accommodating the stacked membrane assembly and achieving the desired dynamic binding capacity is considered to be within the scope of this disclosure. In some embodiments, the intermediate materials 260, 280, the housing 250, the inlet cap 265, and the outlet cap 275 may be formed of a thermoplastic polymer (such as polypropylene, polyethylene, or other polyolefins). Alternatively, one or both of the intermediate materials 260, 280 may be formed of inorganic or metallic materials, provided that the porous intermediate materials 260, 280 do not impede the operation of the chromatographic apparatus.

[0105] The fibrillated heat-treated polymer membrane discs 240 in the stacked membrane assembly 220 can be adhered to the inner wall of the housing 250 by any conventional method (e.g., melt sealing or using a sealant) to prevent flow between the periphery of the fibrillated heat-treated polymer membrane discs 240 and the housing 250. Inlet caps 265 and outlet caps 275 can be sealed to the housing 250 by a similar or identical process. Inlet caps 265 and outlet caps 275 include an inlet 290 and an outlet 285, respectively, to allow the aqueous mixture to flow through the affinity chromatography device 200. Specifically, inlet cap 265 allows fluid of the aqueous mixture to flow into the housing 250, and outlet cap 275 allows fluid of the aqueous mixture to flow out of the housing 250. In use, the aqueous mixture flows sequentially through intermediate material 260, through the fibrillated heat-treated polymer membrane discs 240 forming the stacked membrane assembly 220, and through intermediate material 280. When an aqueous mixture passes through the chromatographic apparatus 200, the affinity ligands reversibly bind to the target molecule, thereby effectively removing it from the aqueous mixture. The target molecule can be removed from the affinity ligands, for example, by passing a liquid with a lower pH value through the apparatus, as is known to those skilled in the art.

[0106] There is no particular limitation on the total number of fibrillated heat-treated polymer membranes present in a stacked membrane assembly, depending on the desired end use and / or the desired mass transport flow rate within the membrane assembly. A stacked membrane assembly may include 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 (or more) layers of polymer membranes. It should be understood that there may be hundreds or even thousands of polymer membrane layers in a stacked membrane assembly. Furthermore, the fibrillated heat-treated polymer membranes present in the stacked membrane assembly may have a thickness of about 1 micrometer to about 10,000 micrometers, about 100 micrometers to about 5,000 micrometers, about 500 micrometers to about 3,000 micrometers, or about 650 micrometers to about 1,000 micrometers. As used herein, the term "thickness" refers to the direction of the fibrillated heat-treated polymer membrane perpendicular to its length-area.

[0107] Both the fibrillated heat-treated polymer films in wound film assemblies and stacked film assemblies contain spherical inorganic particles, or particles with a spherical structure. As used herein, the term "spherical" refers to an inorganic particle having a circular or nearly circular shape, wherein the distance from the center of the inorganic particle to any point on the outer edge of the particle is the same or nearly the same. In some embodiments, the spherical inorganic particles have a particle size distribution of D90 / D10 less than or equal to 3, less than or equal to 2.5, less than or equal to 2, less than or equal to 1.5, or less than or equal to 1. The nominal particle size of the spherical inorganic particles can be about 5 micrometers, about 10 micrometers, about 15 micrometers, about 20 micrometers, and combinations and blends thereof. In some embodiments, the spherical inorganic particles are polydisperse.

[0108] In some embodiments, the protofibrillated heat-treated polymer membrane includes more than one nominal particle size and / or more than one type of spherical inorganic particles. The protofibrous heat-treated polymer film may comprise from about 10% by mass of the protofibrous heat-treated polymer film to about 90% by mass of spherical inorganic particles, from about 15% by mass of the protofibrous heat-treated polymer film to about 85% by mass of spherical inorganic particles, from about 20% by mass of the protofibrous heat-treated polymer film to about 80% by mass of spherical inorganic particles, from about 30% by mass of the protofibrous heat-treated polymer film to about 70% by mass of spherical inorganic particles, from about 35% by mass of the protofibrous heat-treated polymer film to about 65% by mass of spherical inorganic particles, from about 40% by mass of the protofibrous heat-treated polymer film to about 60% by mass of spherical inorganic particles, from about 45% by mass of the protofibrous heat-treated polymer film to about 55% by mass of spherical inorganic particles, or from about 50% by mass of the protofibrous heat-treated polymer film to about 50% by mass of spherical inorganic particles. Non-limiting examples of suitable inorganic particles include silica, zeolite, hydroxyapatite, metal oxides, and combinations thereof. Furthermore, the inorganic particles can be solid or porous. Additionally, the hydraulic permeability of the affinity chromatography apparatus described herein is approximately 200 (X 10⁻⁶). -12 cm 2 ) to approximately 700 (X 10 -12 cm 2 ), approximately 250 (X 10) -12 cm 2 ) to approximately 700 (X 10 -12 cm 2 Approximately 300 (X 10) -12 cm 2 ) to approximately 650 (X 10 -12 cm 2 ), approximately 350 (X 10) -12 cm 2 ) to approximately 600 (X 10 -12 cm 2 ), approximately 400 (X 10) -12 cm 2 ) to approximately 600 (X 10 -12 cm 2 ), approximately 400 (X 10) -12 cm 2 ) to approximately 550 (X 10 -12 cm 2 ), approximately 450 (X 10) -12 cm 2 ) to approximately 550 (X 10 -12 cm 2Approximately 300 (X 10) -12 cm 2 ) to approximately 500 (X 10 -12 cm 2 ), approximately 350 (X 10) -12 cm 2 ) to approximately 500 (X 10 -12 cm 2 ), or approximately 400 (X 10) -12 cm 2 ) to approximately 500 (X 10 -12 cm 2 ).

[0109] In at least one embodiment, the protofibrotic heat-treated polymer film comprises a blend of spherical inorganic particles with different nominal particle sizes. For example, the protofibrotic heat-treated polymer film may comprise a 90:10 mixture of the same or different spherical inorganic particles with a first nominal particle size (e.g., 5 micrometers) and a second nominal particle size (e.g., 20 micrometers). The blend of spherical inorganic particles in the protofibrotic heat-treated polymer film can be any blend, for example, a blend of 10:90 to 90:10, a blend of 30:70 to 70:30, a blend of 60:40 to 40:60, a blend of 25:75 to 75:25, a blend of 20:80 to 80:20, or a blend of 50:50. In one embodiment, the spherical inorganic particles comprise 10-micrometer spherical particles and 20-micrometer spherical particles in a blending ratio of 10:90 to 90:10. In another embodiment, the spherical inorganic particles include 5-micrometer spherical particles and 10-micrometer spherical particles, blended in a ratio of 10:90 to 90:10. In another embodiment, the spherical inorganic particles include 5-micrometer spherical particles and 20-micrometer spherical particles, blended in a ratio of 10:90 to 90:10.

[0110] In some embodiments, the affinity ligand is covalently bonded to spherical inorganic particles. In another embodiment, the affinity ligand is covalently bonded to a protofibrotic heat-treated polymer membrane. In yet another embodiment, the affinity ligand may bind to both the polymer membrane and the spherical inorganic particles. The affinity ligand may be a protein, antibody, or polysaccharide that reversibly binds to a target molecule. In one embodiment, the affinity ligand is a protein that reversibly binds to, for example, the Fc region of an antibody, an antibody fragment, an Fc fusion protein, or an antibody / drug conjugate. In another embodiment, the affinity ligand is an antibody, L protein, or polysaccharide that reversibly binds to a protein or protein fragment it is specific to. Exemplary affinity ligands for affinity chromatography apparatus include, but are not limited to, protein A, protein G, protein L, human Fc receptor proteins, antibodies that specifically bind to other proteins, and heparin. The affinity ligand may be natural, recombinant, or synthetic. In another embodiment, the affinity ligand is a metal affinity ligand that reversibly binds to a histidine-tagged protein. In another embodiment, the affinity ligand may be an antibody or polysaccharide that reversibly binds to the viral vector to which it is specific.

[0111] In at least one embodiment, the protofibrotic heat-treated polymer membrane is an expanded polytetrafluoroethylene (ePTFE) membrane. Expanded polytetrafluoroethylene (ePTFE) membranes prepared according to the methods described in U.S. Patent No. 7,306,729 to Bacino et al., U.S. Patent No. 3,953,566 to Gore, U.S. Patent No. 5,476,589 to Bacino, or U.S. Patent No. 5,183,545 to Branca et al. are applicable herein. Furthermore, the fluoropolymer membrane can be made hydrophilic (e.g., water-wettable) by methods known in the art, such as, but not limited to, the methods disclosed in U.S. Patent No. 4,113,912 to Okita et al., which effectively bind to ligands in coatings, such as applying the membrane as described in U.S. Patent Nos. 5,897,955 to Drumheller, U.S. Patent Nos. 5,914,182 to Drumheller, or U.S. Patent No. 8,591,932 to the polymer membrane.

[0112] The heat-treated fibrillated fluoropolymer membrane may also include polymer materials containing functionalized tetrafluoroethylene (TFE) copolymer membranes, wherein the functionalized TFE copolymer materials include functionalized copolymers of TFE and PSVE (perfluorosulfonyl vinyl ether), or TFE with another suitable functional monomer, such as, but not limited to, vinylidene fluoride (VDF), vinyl acetate, or vinyl alcohol.

[0113] It should be understood that throughout this application, the term "PTFE" is used for convenience and means not only polytetrafluoroethylene, but also expanded PTFE, expanded modified PTFE, and expanded copolymers of PTFE, such as those described in U.S. Patent No. 5,708,044 to Branca, U.S. Patent No. 6,541,589 to Baillie, U.S. Patent No. 7,531,611 to Sabol et al., U.S. Patent No. 8,637,144 to Ford, and U.S. Patent No. 9,139,669 to Xu et al.

[0114] The intermediate material can be a fluoropolymer film or a non-fluoropolymer film (e.g., polyethylene, expanded polyethylene, or other polyolefin films). Furthermore, the intermediate film can be porous. In some embodiments, the intermediate film is a thermoplastic or thermosetting polymer film.

[0115] Advantageously, the chromatographic apparatus can be used multiple times. Furthermore, after each separation process or multiple separation processes, the chromatographic apparatus can be cleaned with a cleaning solution (such as sodium hydroxide, phosphoric acid, citric acid, ethanol, etc.) and reused.

[0116] The affinity chromatography apparatus described herein exhibits a dynamic binding capacity (DBC) greater than 35 mg / ml at a residence time of 20 seconds. Furthermore, it possesses cycle durability of at least 100 cycles at an operating pressure not exceeding 0.3 MPa. Moreover, the apparatus can be used multiple times without losing substantial dynamic binding capacity. Specifically, after each separation process, the apparatus can be cleaned with a cleaning solution (such as sodium hydroxide) and reused. Although embodiments of the wound membrane assembly 110 and the stacked membrane assembly 220 are described herein, it should be understood that any number of fibrillated heat-treated polymer membranes, as well as any and all combinations thereof, including the type of fibrillated polymer membranes, the type and size of the spherical inorganic particles, and the orientation of the fibrillated heat-treated polymer membranes within membrane assemblies 110 and 220, are within the scope of this disclosure. Additionally, some or all of the fibrillated polymer membranes may differ from each other in composition, thickness, permeability, etc.

[0117] The chromatographic apparatus and its components described herein can be manufactured using various methods. In some embodiments, injection molding can be used to manufacture the chromatographic components provided herein. Other suitable methods may include, but are not limited to, extrusion, compression molding, solvent casting, and combinations thereof. Embodiments employing two fibrillated heat-treated polymer membranes co-expanded to produce composite membrane modules are also considered to be within the scope of this disclosure. Such composite membrane modules may comprise two (or more) fibrillated heat-treated polymer membranes that may be co-extruded or integrated together.

[0118] In some implementations, for example Figure 7As roughly described herein, the affinity chromatography apparatus described herein utilizes a dual collector 750 consisting of affinity chromatography apparatuses 700 and 701 arranged in parallel configuration. It should be understood that... Figure 7-12 The affinity chromatography apparatus depicted includes stacked membrane assemblies, wound membrane assemblies, or a combination of stacked and wound membrane assemblies. As shown, an aqueous mixture flows through a main pipe 720 to a distribution element 740. The distribution element 740 is not particularly limited, as long as it distributes the fluid from the aqueous mixture in the main pipe 720 into at least two inlet pipes 760, 761. The split aqueous mixture in the inlet pipes 760, 761 flows into chromatography apparatus 700, 701, where the target molecules are captured. The aqueous solution (i.e., the aqueous mixture excluding the target molecules captured by the affinity ligands) flows out of chromatography apparatus 700, 701 through outlet pipes 780, 781, respectively. The aqueous solutions in the outlet pipes 780, 781 are combined in the distribution element 790 and then recombined into a single aqueous solution flowing into the inlet pipe 795. Figure 8 The image depicts a top view of a dual collector 750 including parallel affinity chromatography devices 700, 701. In some embodiments, the collector 750 is located within a housing 720.

[0119] It should be understood that Figure 7 and Figure 8 The two chromatographic devices 700 and 701 shown are for illustrative purposes only. Multiple (i.e., three or more) chromatographic devices of the parallel configuration described herein can be used in an collector, provided there are similar fluid distribution and resistance between the devices. This similarity between affinity chromatographic devices allows for scalability in device size and performance. The affinity chromatographic devices used in collectors (such as dual collectors, quad collectors, etc.) can be the same or different from each other. Furthermore, affinity chromatographic devices can be used in parallel configuration systems without requiring any changes or additions to collector 750.

[0120] Figure 9 An example of an aggregator that includes multiple chromatographic devices as described herein is a four-aggregator. Figure 10 The image shows a top view of the four-assembly unit 950. (Compared to...) Figure 7 and Figure 8 Similar to the dual collector described herein, the aqueous mixture flows to the distribution element 940. The distribution element 940 divides the fluid of the aqueous mixture into inlet pipes 960, 961, 962, and 963. It should be noted that... Figure 9962 and 963 are concealed behind 961 and 962, respectively. It should be noted that 903 and 904 are concealed behind 901 and 902, respectively. The split aqueous mixtures in inlet pipes 960, 961, 962, and 963 flow into the four-chromatographic apparatus 900, 901, 902, and 903, respectively, where target molecules are captured. The aqueous solutions flow out of the respective chromatographic apparatus 900, 901, 902, and 903 (982 and 983 are not shown) through outlet pipes 980, 981, 982, and 983, respectively. The aqueous solutions in outlet pipes 980 and 981 (982 and 983 are concealed behind 980 and 981 and are therefore not shown) are combined in the distribution element 990 and then recombined into a single aqueous solution. In some embodiments, the four-collector 950 may be located within the housing 920.

[0121] Figure 11 An apparatus 1000 is described, comprising two parallel constructs, collectors 1150 and 1151. Figure 12 The diagram depicts a top view of apparatus 1000. Collectors 1150 and 1151 each include four affinity chromatography devices, wherein... Figure 11 The diagram depicts two affinity chromatography units 1100, 1101 and 1102, 1103 for each of the collectors 1150, 1151. The ability to utilize at least two parallel collectors advantageously allows for increased volumetric capacity while utilizing the chromatographic apparatus described herein. In other words, apparatus 1000 eliminates the need to switch to a larger volumetric chromatography apparatus. Furthermore, for example... Figure 11 The described parallel placement of the collector reduces concerns about overpressurization of the device 1000.

[0122] In use, the aqueous mixture flows to distribution element 1140. Distribution element 1140 divides the aqueous mixture into two distribution pipes 1141 and 1142. The aqueous mixture diverted in distribution pipes 1141 and 1142 is then further divided into inlet pipes by distribution elements 1145 and 1146. Figure 11 In the depicted embodiment, inlet pipes 1160 and 1161 are depicted among the four inlet pipes of collector 1150. Similarly, inlet pipes 1162 and 1163 are depicted among the four inlet pipes of collector 1151. It should be understood that in Figure 11 In this configuration, the remaining inlet pipes 1164, 1165, 1166, and 1167 are concealed behind inlet pipes 1160, 1161, 1162, and 1163. The aqueous mixture in the inlet pipes of the first and second collectors 1150 and 1151 flows into the affinity chromatography apparatus. It should be understood that affinity chromatography apparatuses 1100 and 1101 are depicted in collector 1150, and affinity chromatography apparatuses 1102 and 1103 are depicted in collector 1151. Figure 11The remaining chromatographic devices 1104, 1105, 1106, and 1107 are hidden behind affinity chromatographic devices 1100, 1101, 1102, and 1103. The target molecule is captured in the affinity chromatographic devices.

[0123] Aqueous solutions flow out of each chromatographic unit through outlet pipes. Figure 11 The text describes the outlet pipes 1180 and 1181 of the collector 1150 and the outlet pipes 1182 and 1183 of the collector 1151. It should be understood that the remaining outlet pipes are... Figure 11 The aqueous solution is concealed behind outlet pipes 1180, 1181, 1182, and 1183. The aqueous solution in the outlet pipes is combined in distribution elements 1170 and 1171. The combined solution then flows from distribution elements 1170 and 1171 into collection pipes 1110 and 1111. The aqueous solution in collection pipes 1110 and 1111 is combined in distribution element 1190 and then reassembled into a single aqueous solution. The first collector 1150 and the second collector 1151 can be enclosed within housing 1120.

[0124] In some embodiments, this disclosure relates to a diagnostic device for removing a target substance from a biological sample. The device includes a heat-treated fibrillated polymer membrane containing spherical inorganic particles having a spherical shape and a nominal particle size of about 5 micrometers to about 20 micrometers. The particle size distribution has a D90 / D10 ratio of less than or equal to 3, and at least one of the heat-treated fibrillated polymer membrane and the inorganic particles is covalently bonded to a ligand that reversibly binds to the target substance in the biological sample. The device may further include a fluid inlet and a fluid outlet in fluid communication with the fluid inlet. Additionally, the device may include a housing member containing the fluid inlet, the fluid outlet, and the fibrillated polymer membrane.

[0125] Those skilled in the art will readily understand that various aspects of this disclosure can be implemented by any number of methods and apparatuses configured to perform the desired functions. It should also be noted that the accompanying drawings referenced herein are not necessarily drawn to scale, but may be enlarged to illustrate various aspects of this disclosure, and in this respect, the drawings should not be considered limiting.

[0126] Test methods

[0127] It should be understood that while certain methods and apparatuses are described below, other methods or apparatuses that are determined to be suitable by those skilled in the art may also be used alternatively.

[0128] Methods for measuring membrane permeability

[0129] Membrane permeability was measured using a Gurley Precision Instruments (Troy, New York) meter. The time required for 100 ml of air to permeate through a 1-inch diameter sample was recorded.

[0130] Methods for determining particle size and particle size distribution

[0131] Particle size and particle size distribution data are provided by the manufacturer and measured using Coulter counter technology.

[0132] Methods for determining the melting temperature of PTFE

[0133] The melting temperature of the PTFE within the membrane was characterized using a differential scanning calorimeter (Q2000, TA Instruments, Newcastle, Delaware). Heat flux measurements were performed on a 4 mm diameter membrane sample under nitrogen atmosphere, increasing the temperature from -40 °C to 400 °C at a rate of 10 °C / min. The melting temperature was recorded as the maximum value of the associated endothermic peak.

[0134] Method for determining dynamic binding capacity at 10% penetration

[0135] Insert the chromatographic apparatus into the flow path of the AKTA™ Purification (Cytiva, Marlborough, Massachusetts) liquid chromatography system and perform a single cycle consisting of the following protocol. Table A lists the solutions used; Table B lists the protocol steps for determining the dynamic binding capacity at 10% breakthrough.

[0136] Table A

[0137]

[0138] Table B

[0139]

[0140] Methods for determining liquid permeability

[0141] The liquid permeability of the chromatographic apparatus was determined using Darcy's law. The bed cross-sectional area and bed length of each apparatus were characterized. Solution A was used as the liquid, and its viscosity was characterized. The relationship between the pressure drop across the column and the liquid flow rate was measured on the AKTA™ purified liquid chromatography system.

[0142] Example

[0143] Example 1

[0144] Spherical particles with the particle size and particle size distribution described in Table C were obtained. The porous particles in Table C are substantially the same in terms of chemical and physical properties, such as chemical composition, particle shape, nominal particle porosity, nominal particle pore size, and nominal particle surface area.

[0145] Table C

[0146]

[0147] Porous polytetrafluoroethylene (PTFE) membranes were obtained with 15% by mass of PTFE and 85% by mass of the spherical porous silica particles shown in Table G. These ratios are shown in Table D.

[0148] Table D

[0149]

[0150] Adjacent portions of the porous membrane described in Table D were fixed to a frame and subjected to a heat treatment process at 370°C for 20 seconds. The membrane was then removed while still within the frame and cooled to room temperature. Table E lists the physical properties and permeability of the polymer membrane before and after the heat treatment process.

[0151] Table E

[0152]

[0153] The PTFE melting temperatures of porous membranes A and D before and after heat treatment were characterized. The results are shown in Table F.

[0154] Table F

[0155]

[0156] The porous membranes in Table E, both before and after heat treatment, are used to fabricate affinity chromatography devices. A polypropylene fluid distributor is fixed to one end of a polypropylene cylindrical shell. Porous polypropylene intermediate material is placed inside the shell. As shown in Figure G, the desired number of PTFE membrane layers are stacked on top of the polypropylene intermediate material inside the shell. A second porous polypropylene intermediate material is placed on top of the PTFE membrane layers. A second polypropylene fluid distributor is fixed to the end of the cylindrical shell opposite the first polypropylene fluid distributor. The chromatography device is sealed by heating.

[0157] The intermediate device is then treated in a manner that covalently bonds protein A to the stacked PTFE membrane. This approach represents a typical approach for those skilled in the art and is further described in U.S. Patent Nos. 10,525,376 and 10,526,367 to McManaway et al.

[0158] The affinity chromatography devices manufactured above were tested, and their liquid permeability and dynamic binding capacity at a residence time of twenty (20) seconds were evaluated using the test methods described herein. The performance of each affinity chromatography device is shown in Table G.

[0159] Table G

[0160]

[0161] The relative effects of membrane heat treatment on the liquid permeability and dynamic binding capacity shown in Table G. Figure 5 The information is presented in the form of charts.

[0162] Example 2

[0163] The type A porous membranes in Table E, both before and after heat treatment, are used to manufacture affinity chromatography devices. A polypropylene fluid distributor is fixed to one end of a polypropylene cylindrical shell. Porous polypropylene intermediate material is placed inside the shell. As shown in Figure H, the desired number of PTFE membrane layers are stacked on top of the polypropylene intermediate material inside the shell. A second porous polypropylene intermediate material is placed on top of the PTFE membrane layers. A second polypropylene fluid distributor is fixed to the end of the cylindrical shell opposite the first polypropylene fluid distributor. The chromatography device is sealed by heating.

[0164] As described in Example 1, the intermediate device was then processed in a manner that covalently bonded protein A to the stacked membrane, and then tested to evaluate its liquid permeability and dynamic binding capacity at a residence time of twenty (20) seconds. The performance of each affinity chromatography device is shown in Table H.

[0165] Table H

[0166]

[0167] like Figure 7 and Figure 8 As described, devices I and J are connected in parallel to form device M. Devices K and L are also connected in parallel to form device N.

[0168] The aforementioned affinity chromatography devices M and N were tested, and their liquid permeability and dynamic binding capacity at a residence time of twenty (20) seconds were evaluated using the test methods described herein. The performance of each affinity chromatography device is shown in Table I.

[0169] Table I

[0170]

[0171] The relative effects of membrane heat treatment on the liquid permeability and dynamic binding capacity shown in Table G. Figure 6 It is represented in the form of a chart and can be compared with... Figure 5 The data comparison presented in the text.

[0172] The invention of this application has been described for brevity and in conjunction with specific embodiments above. It will be apparent to those skilled in the art that various modifications and variations can be made to the embodiments described herein without departing from the spirit and scope of the invention. Therefore, the embodiments are intended to cover these modifications and variations of the invention, provided that such modifications and variations are within the scope of the appended claims and their equivalents.

Claims

1. An article comprising: Includes the following affinity chromatography apparatus: Fluid inlet; The fluid outlet is connected to the fluid inlet. A heat-treated fibrillated polymer membrane located between a fluid inlet and a fluid outlet, comprising inorganic particles having a spherical shape and a nominal particle size selected from 5 micrometers, 10 micrometers, 15 micrometers, 20 micrometers and combinations thereof; The shell surrounding the fluid inlet, fluid outlet, and the original fibrous heat-treated polymer membrane. The particle size distribution has a D90 / D10 ratio of less than or equal to 3. In the case of the protofibrotic heat-treated polymer membrane and the inorganic particles, at least one of them has been covalently bonded to an affinity ligand, which reversibly binds to proteins, antibodies, viral vectors, and combinations thereof; The affinity chromatography device described herein has approximately 200 (X10) -12 cm 2 ) to approximately 700 (X10) -12 cm 2 Hydraulic penetration rate The affinity chromatography device described herein has a dynamic binding capacity (DBC) of at least 35 mg / ml at a residence time of 20 seconds, and The fibrillated heat-treated polymer membrane is obtained by subjecting a fibrillated polymer membrane containing inorganic particles to a temperature higher than the melting point of the fibrillated polymer membrane for a specified time period.

2. The article of claim 1, wherein the inorganic particles having a spherical shape are selected from silica, zeolite, hydroxyapatite, metal oxides and combinations thereof.

3. The article of claim 1, wherein the original fiberized heat-treated polymer film comprises expanded polytetrafluoroethylene film, expanded modified polytetrafluoroethylene film, expanded polytetrafluoroethylene copolymer film or expanded polyethylene film.

4. The article of claim 1, wherein the original fibrous polymer membrane is an expanded polytetrafluoroethylene membrane.

5. The article of claim 1, wherein the affinity ligand is selected from protein A, protein G, protein L, human Fc receptor protein, antibody, polysaccharide, and combination thereof.

6. The article of claim 1, wherein the inorganic particles comprise at least a first inorganic particle having a spherical shape and a first nominal particle size, and a second inorganic particle having a spherical shape and a second nominal particle size, the first nominal particle size and the second nominal particle size being different from each other.

7. The article of claim 1, wherein the inorganic particles having a spherical shape comprise a blend of 10-micrometer spherical particles and 20-micrometer spherical particles, and wherein the blend is 10:90 to 90:

10.

8. The article of claim 1, wherein the inorganic particles having a spherical shape comprise a blend of 5-micrometer spherical particles and 10-micrometer spherical particles, and wherein the blend is 10:90 to 90:

10.

9. The article of claim 1, wherein the inorganic particles having a spherical shape comprise a blend of 5-micrometer spherical particles and 20-micrometer spherical particles, and wherein the blend is 10:90 to 90:

10.

10. The article of claim 1, comprising cyclic durability of at least 100 cycles at an operating pressure of less than 0.3 MPa.

11. The article of claim 1, wherein the original fibrous heat-treated polymer film has a wound structure.

12. The article of claim 1, wherein the fibrillated polymer film has a wound structure, a stacked structure, or a combination thereof.

13. The article of claim 1, wherein the inner intermediate material surrounds the outer surface of the core, and wherein the protofibrotic heat-treated polymer film surrounds the inner intermediate material.

14. The article of claim 13, comprising an outer intermediate material surrounding a heat-treated fibrillated polymer membrane.

15. The article of claim 14, wherein the inner intermediate material and the outer intermediate material are selected from porous fluoropolymer membranes, porous non-fluoropolymer membranes, porous nonwoven materials, and porous woven materials.

16. The article of claim 14, wherein at least one of the inner layer intermediate material and the outer layer intermediate material is a polypropylene nonwoven material.

17. An assembly comprising at least two articles of any one of claims 1 to 16 arranged in parallel configuration.

18. The collector of claim 17, wherein the collector is housed in a housing.

19. Use of the article of any one of claims 1 to 16 for separating proteins, antibodies, viral vectors and combinations thereof from a fluid stream.

20. An apparatus comprising a first collector and a second collector constructed in parallel, wherein each of the first collector and the second collector comprises at least two articles of any one of claims 1 to 16.

21. The apparatus of claim 20, wherein the first collector and the second collector are enclosed in a housing.

22. An article comprising: The core located at the center; A protofibrotic heat-treated polymer membrane wrapped around a core, comprising spherical inorganic particles having a spherical shape and a nominal particle size selected from 5 micrometers, 10 micrometers, 15 micrometers, 20 micrometers and combinations thereof; The shell surrounding the core and the original fibrous heat-treated polymer film, The first end cap is located at the first end of the housing, and The second end cap is located at the second end of the housing. The particle size distribution has a D90 / D10 ratio of less than 3. In the case of the protofibrotic heat-treated polymer membrane and the spherical inorganic particles, at least one of them has been covalently bonded to an affinity ligand, which reversibly binds to proteins, antibodies, viral vectors, and combinations thereof; The article described herein has approximately 200 (X10) -12 (cm2) to approximately 700 (X10) -12 Hydraulic penetration rate (cm2) The article described above has a dynamic binding capacity (DBC) of at least 35 mg / ml at a residence time of 20 seconds, and The fibrillated heat-treated polymer membrane is obtained by subjecting a fibrillated polymer membrane containing inorganic particles to a temperature higher than the melting point of the fibrillated polymer membrane for a specified time period.

23. The article of claim 22, wherein the inorganic particles having a spherical shape are selected from silica, zeolite, hydroxyapatite, metal oxides and combinations thereof.

24. The article of claim 22, wherein the original fiberized heat-treated polymer film comprises at least one of expanded polytetrafluoroethylene film, expanded modified polytetrafluoroethylene film, expanded polytetrafluoroethylene copolymer film, or expanded polyethylene film.

25. The article of claim 22, wherein the original fiberized heat-treated polymer film is an original fiberized heat-treated expanded polytetrafluoroethylene film.

26. The article of claim 22, wherein the affinity ligand is selected from protein A, protein G, protein L, human Fc receptor protein, antibody, polysaccharide, and combination thereof.

27. The article of claim 22, wherein the inorganic particles comprise at least a first inorganic particle having a spherical shape and a first nominal particle size, and a second inorganic particle having a spherical shape and a second nominal particle size, the first nominal particle size and the second nominal particle size being different from each other.

28. The article of claim 22, wherein the inorganic particles having a spherical shape comprise a blend of 10-micrometer spherical particles and 20-micrometer spherical particles, and wherein the blend is 10:90 to 90:

10.

29. The article of claim 22, wherein the inorganic particles having a spherical shape comprise a blend of 5-micron spherical particles and 10-micron spherical particles, and wherein the blend is 10:90 to 90:

10.

30. The article of claim 22, wherein the inorganic particles having a spherical shape comprise a blend of 5-micron spherical particles and 20-micron spherical particles, and wherein the blend is 10:90 to 90:

10.

31. The article of claim 22, comprising a cycle durability of at least 100 cycles and an operating pressure of less than 0.3 MPa.

32. The article of claim 22, wherein the inner intermediate material surrounds the outer surface of the core and the original fibrous heat-treated polymer film surrounds the inner intermediate material.

33. The article of claim 32, comprising an outer intermediate material surrounding the original fibrous heat-treated polymer film.

34. The article of claim 33, wherein the inner intermediate material and the outer intermediate material are selected from porous fluoropolymer membranes, porous non-fluoropolymer membranes, porous nonwoven materials and porous woven materials.

35. The article of claim 33, wherein at least one of the inner layer intermediate material and the outer layer intermediate material is a polypropylene nonwoven material.

36. An assembly comprising at least two articles of any one of claims 22 to 35 arranged in parallel configuration.

37. The collector of claim 36, wherein the collector is enclosed in a housing.

38. An apparatus comprising a first collector and a second collector constructed in parallel, wherein each of the first collector and the second collector comprises at least two articles of any one of claims 22 to 35.

39. The apparatus of claim 38, wherein the first collector and the second collector are enclosed in a housing.

40. Use of the article of any one of claims 22 to 35 for separating proteins, antibodies, viral vectors and combinations thereof from a fluid stream.

41. An article comprising: Includes the following affinity chromatography apparatus: Shell components; Allowing fluid to flow into the inlet of the housing; The outlet of the housing allows the fluid to flow out. and A stacked membrane assembly disposed within the housing between a fluid inlet and a fluid outlet, the stacked membrane assembly comprising: A two- or multi-layer stacked fibrillated heat-treated polymer membrane, each layer of which contains inorganic particles having a spherical shape and a blended nominal particle size selected from 5 micrometers, 10 micrometers, 15 micrometers, 20 micrometers, and combinations thereof. The particle size distribution has a D90 / D10 ratio of less than 3, and In the case of the protofibrotic heat-treated polymer membrane and the spherical inorganic particles, at least one of them has been covalently bonded to an affinity ligand, which reversibly binds to proteins, antibodies, viral vectors, and combinations thereof; The affinity chromatography device described herein has approximately 200 (X10) -12 (cm2) to approximately 700 (X10) -12 Hydraulic penetration rate (cm2) The affinity chromatography device described herein has a dynamic binding capacity (DBC) of at least 35 mg / ml at a residence time of 20 seconds, and Each of the aforementioned fibrillated heat-treated polymer membranes is obtained by subjecting a fibrillated polymer membrane containing inorganic particles to a temperature above the melting point of the fibrillated polymer membrane for a specified time period.

42. The article of claim 41, comprising first and second fluid dispensers located at opposite ends of the housing.

43. The article of claim 41, wherein the inorganic particles having a spherical shape are selected from silica, zeolite, hydroxyapatite, metal oxides and combinations thereof.

44. The article of claim 41, wherein the original fiberized heat-treated polymer film comprises expanded polytetrafluoroethylene film, expanded modified polytetrafluoroethylene film, expanded polytetrafluoroethylene copolymer film, or expanded polyethylene film.

45. The article of claim 41, wherein the original fiberized heat-treated polymer film is an original fiberized heat-treated expanded polytetrafluoroethylene film.

46. ​​The article of claim 41, wherein the affinity ligand is selected from protein A, protein G, protein L, human Fc receptor protein, antibody, polysaccharide, and combination thereof.

47. The article of claim 41, wherein the inorganic particles comprise at least a first inorganic particle having a spherical shape and a first nominal particle size, and a second inorganic particle having a spherical shape and a second nominal particle size, the first nominal particle size and the second nominal particle size being different from each other.

48. The article of claim 41, wherein the inorganic particles having a spherical shape comprise a blend of 10-micrometer spherical particles and 20-micrometer spherical particles, and wherein the blend is 90:10 to 10:

90.

49. The article of claim 41, wherein the inorganic particles having a spherical shape comprise a blend of 5-micrometer spherical particles and 10-micrometer spherical particles, and wherein the blend is 10:90 to 90:

10.

50. The article of claim 41, wherein the inorganic particles having a spherical shape comprise a blend of 5-micrometer spherical particles and 20-micrometer spherical particles, and wherein the blend is 10:90 to 90:

10.

51. The article of claim 41, comprising cyclic durability of at least 100 cycles at an operating pressure of less than 0.3 MPa.

52. The article of claim 41, wherein at least one first intermediate material is located on a first side of the stacked film assembly and a second intermediate material is located on a second side of the stacked film assembly, the second side being opposite to the first side.

53. The article of claim 52, wherein the first intermediate material and the second intermediate material are selected from porous fluoropolymer membranes, porous non-fluoropolymer membranes, porous nonwoven materials and porous woven materials.

54. The article of claim 53, wherein at least one of the first intermediate material and the second intermediate material is a polypropylene nonwoven material.

55. An assembler comprising at least two articles of any one of claims 41 to 54 arranged in parallel configuration.

56. The collector of claim 55, wherein the collector is enclosed in a housing.

57. An apparatus comprising a first collector and a second collector constructed in parallel, wherein each of the first collector and the second collector comprises at least two articles of any one of claims 41 to 54.

58. The apparatus of claim 57, wherein the first collector and the second collector are enclosed in a housing.

59. An article comprising: A diagnostic device comprising: A fibrillated heat-treated polymer membrane is located between a fluid inlet and a fluid outlet, and contains inorganic particles with a spherical shape and a nominal particle size selected from 5 micrometers, 10 micrometers, 15 micrometers, 20 micrometers, and combinations thereof. The particle size distribution has a D90 / D10 ratio of less than or equal to 3. In this embodiment, at least one of the fibrillated heat-treated polymer membrane and the inorganic particles is covalently bonded to a ligand, which reversibly binds to proteins, antibodies, viral vectors, or combinations thereof within the biological sample. The article described herein has approximately 200 (X10) -12 cm 2 ) to approximately 700 (X10) -12 cm 2 Hydraulic penetration rate The article described above has a dynamic binding capacity (DBC) of at least 35 mg / ml at a residence time of 20 seconds, and The fibrillated heat-treated polymer membrane is obtained by subjecting a fibrillated polymer membrane containing inorganic particles to a temperature higher than the melting point of the fibrillated polymer membrane for a specified time period.

60. The article of claim 59, comprising a fluid inlet and a fluid outlet fluidly connected to the fluid inlet.

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