EP4658394A1

EP4658394A1 - Dry fibrillated membranes and affinity chromatography devices containing the same

Info

Publication number: EP4658394A1
Application number: EP24710957.2A
Authority: EP
Inventors: Jared M. CLINGER; Michael C. MCMANAWAY
Original assignee: WL Gore and Associates Inc
Current assignee: WL Gore and Associates Inc
Priority date: 2023-01-31
Filing date: 2024-01-31
Publication date: 2025-12-10
Also published as: JP2026507742A; KR20250141749A; WO2024163661A1; CN120752083A

Abstract

The present disclosure is directed to affinity chromatography devices that include a dry fibrillated polymer membrane having a moisture content less than or equal to 60% by mass of the dry fibrillated membrane. In forming the dry fibrillated polymer membrane, a non-dry fibrillated polymer membrane may be dried through lyophilization. Dry affinity chromatography devices may improve the stability or viability of a target molecule being purified, decrease shipping risks (for example, reduces risk in the event of an accidental rupture or leak due to a drop), and reduce storage risk (for example, reduces risk of microbial growth during storage). The affinity chromatography device and/or the dry fibrillated polymer membrane may be treated with ethylene oxide, gamma irradiation, or x-ray irradiation. In some embodiments, the treatment sterilizes the chromatography device or dry fibrillated polymer membrane. Prior to use the dry affinity chromatography devices can be wetted with bioprocessing fluids.

Description

DRY FIBRILLATED MEMBRANES AND AFFINITY CHROMATOGRAPHY DEVICES CONTAINING THE SAME

FIELD

[001] The present disclosure relates generally to affinity chromatography, and more specifically, to dry fibri Hated membranes containing therein inorganic particles and affinity chromatography devices that include a dry fibrillated polymer membrane that enables the separation of a targeted molecule from an aqueous mixture.

BACKGROUND

[002] Chromatographic methods generally are used to separate and/or purify molecules of interest such as proteins, nucleic acids, and polysaccharides from a mixture. Affinity chromatography specifically involves passing the mixture over a matrix having a ligand specific (/.e., a specific binding partner) for the molecule of interest bound to it. Upon contacting the ligand, the molecule of interest is bound to the matrix and is therefore retained from the mixture. Affinity chromatography provides some advantages over other types of chromatography. For example, affinity chromatography provides a purification method that can isolate a target protein from a mixture of the target protein and other biomolecules in a single step in high yield.

[003] Despite the advantages of current affinity chromatography devices, there exists a need in the art for a chromatography device that can be used at shorter residence times than conventional devices while providing the same binding capacity or better binding capacities than current offerings, that is re-useable, ready-to-use and/or ready-to-sterilize and having reduced shipping and/or storage risk.

SUMMARY

[004] According to one aspect (“Aspect 1”), a composite membrane for an affinity chromatography device includes a dry fibrillated polymer membrane having therein inorganic particles where at least one of the inorganic particles and dry fibrillated polymer membrane have covalently bonded thereto an affinity ligand that reversibly binds to a targeted molecule and where the dry fibrillated polymer membrane has a moisture content of less than or equal to 60% by mass of the dry fibrillated membrane. [005] According to another aspect (Aspect 2), further to Aspect 1 , the inorganic particles are selected from spherical particles, non-spherical particles, and combinations thereof.

[006] According to another aspect (Aspect 3), further to Aspect 1 or Aspect 2, the inorganic particles have a spherical shape and a nominal particle size from about 5 microns to about 20 microns.

[007] According to another aspect (Aspect 4), further to any one of Aspects 1 to

3, the inorganic particles have a particle size distribution has a D90/D10 less than or equal to 3.

[008] According to another aspect (Aspect 5), further to any one of Aspects 1 to

4, the dry fibril lated polymer membrane includes an expanded polytetrafluoroethylene membrane, an expanded modified polytetrafluoroethylene membrane, an expanded tetrafluoroethylene copolymer membrane, or an expanded polyethylene membrane.

[009] According to another aspect (Aspect 6), further to any one of Aspects 1 to

5, the affinity ligand is selected from Protein A, Protein G, Protein L, human Fc receptor protein, antibodies, polysaccharides, oligosaccharides, oligonucleotides, and combinations thereof.

[0010] According to another aspect (Aspect 7), further to any one of Aspects 1 to

6, the targeted molecule comprises a protein, antibody, viral vector, nucleic acid or combinations thereof.

[0011] According to another aspect (Aspect 8), further to any one of Aspects 1 to

7, the dry f ibril lated polymer membrane has a wound configuration.

[0012] According to another aspect (Aspect 9), further to any one of Aspects 1 to

8, the dry fibril lated polymer membrane has a stacked configuration.

[0013] According to another aspect (Aspect 10), further to any one of Aspects 1 to 9, the dry fibril lated polymer membrane is treated for sterilization with ethylene oxide, gamma irradiation, or x-ray irradiation.

[0014] According to one aspect (“Aspect 11”), an affinity chromatography device includes a fluid inlet, a fluid outlet fluidly connected to the fluid inlet, a dry fibri Hated polymer membrane positioned between the fluid inlet and the fluid outlet and containing therein at least one inorganic particle where at least one of (1 ) the dry fibril lated polymer membrane has a moisture content less than or equal to 60% by mass of the dry f ibril lated polymer membrane prior to drying, or (2) the affinity chromatography device has a moisture content less than or equal to 60% by mass of the affinity chromatography device, and where at least one of the dry fibrillated polymer membrane and the inorganic particles has covalently bonded thereto an affinity ligand that reversibly binds to a targeted molecule.

[0015] According to another aspect (Aspect 12), further to Aspect 11 , including a housing encompassing the fluid inlet, the fluid outlet, and the dry fibrillated polymer membrane.

[0016] According to another aspect (Aspect 13), further to Aspect 11 or Aspect 12, two or more of the dry fibrillated membrane is in the form of a dry stacked membrane assembly positioned between the fluid inlet and fluid outlet.

[0017] According to another aspect (Aspect 14), further to Aspect 11 or Aspect 12, the dry fibrillated membrane has a wound membrane configuration including the dry fibrillated polymer wound around a core.

[0018] According to another aspect (Aspect 15), further to any one of Aspects 11 to 14, the targeted molecule comprises a protein, antibody, viral vector, nucleic acid or combinations thereof.

[0019] According to another aspect (Aspect 16), further to any one of Aspects 11 to 15, the inorganic particles are selected from spherical particles, non-spherical particles, and combinations thereof.

[0020] According to another aspect (Aspect 17), further to any one of Aspects 11 to 16, the dry fibrillated polymer membrane comprises an expanded polytetrafluoroethylene membrane, an expanded modified polytetrafluoroethylene membrane, an expanded tetrafluoroethylene copolymer membrane, or an expanded polyethylene membrane.

[0021] According to another aspect (Aspect 18), further to any one of Aspects 11 to 17, the affinity ligand is selected from Protein A, Protein G, Protein L, human Fc receptor protein, antibodies, polysaccharides, oligosaccharides, and combinations thereof. [0022] According to another aspect (Aspect 19), further to any one of Aspects 11 to 18, at least one of the dry fibrillated polymer membrane and the affinity chromatography device is treated with ethylene oxide, gamma irradiation or x-ray irradiation.

[0023] According to another aspect (Aspect 20), further to any one of Aspects 11 to 19, the affinity chromatography device is configured to achieve a dynamic binding capacity (DBC) of at least 35 mg/ml at a residence time of 20 seconds.

[0024] According to another aspect (Aspect 21 ), further to any one of Aspects 11 to 20, the affinity chromatography device is configured to achieve a hydraulic permeability from about 100 (X 10'¹² cm²) to about 2000 (X 10'¹² cm²).

[0025] According to another aspect (Aspect 22), further to any one of Aspects 11 to 21 , the affinity chromatography device is configured to achieve an elution volume from 100 mAU to 100 mAU of 1 column volume (CV) to 6 column volumes (CV).

[0026] According to another aspect (Aspect 23), further to any one of Aspects 11 to 22, the affinity chromatography device is configured to achieve a cycling durability of at least 100 cycles at an operating pressure less than 0.3 MPa.

[0027] According to another aspect (Aspect 24), further to any one of Aspects 11 to 23, use of the article to separate the targeted molecule from a fluid stream.

[0028] According to another aspect (Aspect 25), a manifold including at least two of the affinity chromatography devices of any one of Aspects 11 to 23 are arranged in a parallel configuration.

[0029] According to another aspect (Aspect 26), further to Aspect 25, the manifold is enclosed within a housing.

[0030] According to another aspect (Aspect 27), a device including a first manifold and a second manifold are arranged in a parallel configuration where each of the first manifold and the second manifold includes at least two of the affinity chromatography devices of any one of Aspects 11 to 23.

[0031] According to another aspect (Aspect 28), further to Aspect 27, the first manifold and the second manifold are enclosed within a housing.

[0032] According to another aspect (Aspect 29), a method of isolating a targeted molecule includes (1 ) providing a chromatography device that includes an inlet, an outlet, and a dry fibri Hated polymer membrane with a moisture content less than or equal to 60% by mass of the dry fibril lated membrane, where the dry fibrillated polymer membrane is positioned between the inlet and the outlet and contains therein at least one inorganic particle, (2) wetting the dry fibrillated polymer membrane to form a wet polymer membrane, (3) adding an aqueous mixture containing therein a target molecule to the inlet, (4) passing the aqueous mixture through the wet fibrillated polymer membrane where the target molecule is bound to an affinity ligand, and (5) binding the target molecule to an affinity ligand, where at least one of the dry fibrillated polymer membrane and the inorganic particle has thereon the affinity ligand.

[0033] According to another aspect (Aspect 30), further to Aspect 29, including treating the dry fibrillated polymer membrane with ethylene oxide to sterilize the dry fibrillated polymer membrane.

[0034] According to another aspect (Aspect 31 ), further to Aspect 30, where the treating is conducted in accordance with the Method for Ethylene Oxide (EO) Treatment of Membranes set forth in the Test Method section.

[0035] According to another aspect (Aspect 32), further to Aspect 29, including treating the chromatography device with ethylene oxide, gamma irradiation, or x-ray irradiation.

[0036] According to another aspect (Aspect 33), further to Aspect 29, including treating the chromatography device with ethylene oxide, gamma irradiation, or x-ray irradiation to sterilize the chromatography device.

[0037] According to another aspect (Aspect 34), further to Aspect 29, including treating a non-dry fibrillated membrane to covalently bond an affinity ligand to one or both of the non-dry fibrillated polymer membrane and inorganic particle.

[0038] According to another aspect (Aspect 35), further to Aspect 29, including characterizing the moisture content of the dry fibrillated polymer membrane.

[0039] According to another aspect (Aspect 36), further to Aspect 35, where the characterizing is conducted according to the Method for Determining Moisture Content of Membranes set forth in the Test Methods section.

[0040] According to another aspect (Aspect 37), further to Aspect 29, including characterizing the moisture content of the chromatography device. [0041] According to another aspect (Aspect 38), further to Aspect 29, including drying a non-dry fibri Hated membrane through lyophilization to form the dry fibril lated polymer membrane.

[0042] According to another aspect (Aspect 39), further to Aspect 38, where the drying is conducted according to the Method of Lyophilization of Membranes set forth in the Test Methods section.

[0043] According to another aspect (Aspect 40), further to Aspect 29, including determining the dynamic binding capacity at 10% breakthrough.

[0044] According to another aspect (Aspect 41 ), further to Aspect 40, where the determining is conducted according to the Method for Determining the Dynamic Binding Capacity at 10% Breakthrough for Device Articles set forth in the Test Methods section.

[0045] According to another aspect (Aspect 42), further to Aspect 29, including initially wetting the dry chromatography device.

[0046] According to another aspect (Aspect 43), further to Aspect 42, where the initially wetting is conducted according to the Method for Initial Wetting of Dry Device Articles set forth in the Test Methods section.

[0047] According to another aspect (Aspect 44), further to Aspect 29, including determining an elution volume for the chromatography device.

[0048] According to another aspect (Aspect 45), further to Aspect 44, where the determining is conducted according to the Method for Determining Elution Volume from 100m AU to 100mAU for Device Articles set forth in the Test Methods section.

[0049] According to another aspect (Aspect 46), further to Aspect 29, including determining the dynamic binding capacity at 10% breakthrough for the chromatograph device.

[0050] According to another aspect (Aspect 47), further to Aspect 46, where the determining is conducted according to the Method for Determining the Dynamic Binding Capacity at 10% Breakthrough for Device Articles set forth in the Test Methods section.

[0051] According to another aspect (Aspect 48), further to Aspect 29, where the chromatography device is an affinity chromatography device. [0052] According to another aspect (Aspect 49), further to Aspect 29, the affinity ligand is selected from Protein A, Protein G, Protein L, human Fc receptor protein, antibodies, polysaccharides, oligosaccharides, and combinations thereof.

[0053] According to another aspect (Aspect 50), further to Aspect 29, the inorganic particle are selected from spherical particles, non-spherical particles, and combinations thereof.

[0054] According to another aspect (Aspect 51 ), further to Aspect 29, the dry fibrillated polymer includes an expanded polytetrafluoroethylene membrane, an expanded modified polytetrafluoroethylene membrane, an expanded tetrafluoroethylene copolymer membrane, or an expanded polyethylene membrane.

[0055] According to another aspect (Aspect 52), an affinity chromatography device includes a housing member, an inlet to permit fluid flow into the housing member, an outlet to permit fluid flow out of the housing member and fluidly connected to the inlet, a stacked membrane assembly positioned within the housing member between the fluid inlet and fluid outlet where the stacked membrane includes two or more dry fibrillated polymer membranes in a stacked configuration, each of the dry fibrillated polymer membrane containing therein inorganic particles, where the dry fibrillated polymer membrane has a moisture content of less than or equal to 60% by mass of the dry fibrillated polymer membrane, and at least one of the dry fibrillated polymer membrane and the inorganic particles has covalently bonded thereto an affinity ligand that reversibly binds to a targeted molecule.

[0056] According to another aspect (Aspect 53), further to Aspect 52, the targeted molecule includes a protein, antibody, viral vector, nucleic acid or combinations thereof.

[0057] According to another aspect (Aspect 54), further to Aspect 51 or Aspect 52, the inorganic particles are selected from spherical particles, non-spherical particles, and combinations thereof.

[0058] According to another aspect (Aspect 55), further to any one of Aspects 52 to 54, the inorganic particles have a spherical shape and a nominal particle size from about 5 microns to about 20 microns. [0059] According to another aspect (Aspect 56), further to any one of Aspects 52 to 55, a particle size distribution has a D90/D10 less than or equal to 3.

[0060] According to another aspect (Aspect 57), further to any one of Aspects 52 to 56, the dry fibrillated polymer membrane comprises an expanded polytetrafluoroethylene membrane, an expanded modified polytetrafluoroethylene membrane, an expanded tetrafluoroethylene copolymer membrane, or an expanded polyethylene membrane.

[0061] According to another aspect (Aspect 58), further to any one of Aspects 52 to 57, the affinity ligand is selected from Protein A, Protein G, Protein L, human Fc receptor protein, antibodies, polysaccharides, oligosaccharides, oligonucleotides and combinations thereof.

[0062] According to another aspect (Aspect 59), further to any one of Aspects 52 to 58, the device is treated with ethylene oxide, gamma irradiation, or x-ray irradiation.

[0063] According to another aspect (Aspect 60), further to any one of Aspects 52 to 59, the dry fibrillated polymer membrane has a stacked configuration with two sides and a plurality of edges of the stacked membrane assembly, and wherein the device further comprises a material sealing the edges of the stacked membrane assembly.

[0064] The foregoing Examples are just that and should not be read to limit or otherwise narrow the scope of any of the inventive concepts otherwise provided by the instant disclosure. While multiple examples are disclosed, still other embodiments will become apparent to those skilled in the art from the following detailed description, which shows and describes illustrative examples. Accordingly, the drawings and detailed description are to be regarded as illustrative in nature rather than restrictive in nature.

BRIEF DESCRIPTION OF THE DRAWINGS

[0065] The accompanying drawings are included to provide a further understanding of the disclosure and are incorporated in and constitute a part of this specification, illustrate embodiments, and together with the description serve to explain the principles of the disclosure. [0066] FIG. 1 is an exploded view of a chromatography device containing a wound membrane assembly including a dry fibrillated polymer membrane containing therein inorganic particles in accordance with at least one embodiment;

[0067] FIG. 2 is a cross-sectional view of a chromatography device containing a wound membrane assembly depicting a dry fibrillated polymer membrane, an outer flow channel, and an inner flow channel in accordance with at least one embodiment;

[0068] FIG. 3 is an exploded view of a chromatography device containing a stacked membrane assembly including a dry fibrillated polymer membrane containing therein inorganic particles in accordance with at least one embodiment;

[0069] FIG. 4A is a cross-sectional view of a chromatography device containing a stacked membrane assembly including a dry fibrillated polymer membrane containing therein inorganic particles in accordance with at least one embodiment;

[0070] FIG. 4B is an exploded view of the chromatography device of FIG. 4A in accordance with at least one embodiment;

[0071] FIG. 5 is a side view of a manifold having a plurality of chromatography devices in a parallel configuration, where each chromatography device contains a stacked membrane assembly including a dry fibrillated polymer membrane containing therein inorganic particles in accordance with at least one embodiment;

[0072] FIG. 6 is a schematic illustration of a side view of two manifolds in a parallel configuration in accordance with at least one embodiment;

[0073] FIG. 7 is a side view of a manifold having a plurality of chromatography devices in a parallel configuration, each chromatography device containing a wound membrane assembly that includes a dry fibrillated polymer membrane containing therein inorganic particles in accordance with at least one embodiment;

[0074] FIG. 8 is a side view of two manifolds containing a wound dry fibrillated polymer membrane assembly in a parallel configuration in accordance with at least one embodiment;

[0075] FIG. 9 is a UV absorbance at 280 nm chromatogram associated with the characterization of Device T in accordance with at least one embodiment;

[0076] FIG. 10A is a schematic illustration of a multi-well plate in accordance with at least one embodiment; and [0077] FIG. 10B is a is a schematic illustration of a portion of the multi-well plate depicted in FIG. 10A showing a portion of a stacked membrane assembly positioned on a porous substrate in accordance with at least one embodiment.

DETAILED DESCRIPTION

[0078] Persons skilled in the art will readily appreciate that the accompanying figures referred to herein are not necessarily drawn to scale but may be exaggerated to illustrate various aspects of the present disclosure, and in that regard, the figures should not be construed as limiting.

[0079] As used herein, the term “on” is meant to denote an element, such as a polymer membrane, is directly on another element or intervening elements may also be present.

[0080] As used herein, the phrases “spherical particles”, “spherical inorganic particles”, and “inorganic particles having a spherical shape” may be used interchangeably.

[0081] As used herein, the term “fibrillated polymer membrane” is meant to refer to the presence of fibrils in the polymer membrane, such as, for example, a polymer membrane having a microstructure that is characterized by nodes, fibrils, and voids where the nodes are interconnected by the fibrils and the voids are the spaces positioned between the nodes and fibrils.

[0082] The terms “fibrillated polymer membrane”, “polymer membrane”, and “membrane” may be used interchangeably herein.

[0083] As used herein, the term “spiral wound membrane assembly” is meant to include both the dry fibrillated polymer membrane alone and the dry fibrillated polymer membrane with one or more intermediate material(s) in a spiral configuration.

[0084] As used herein, the term “stacked membrane assembly” is meant to include both the dry fibrillated polymer membrane alone and the dry fibrillated polymer membrane with one or more intermediate material(s) in a stacked configuration.

[0085] As used herein, the terms “dry affinity chromatography device”, “dry chromatography device”, “dry device”, and “dry device article” may be used interchangeably. [0086] The term “wet fibril lated polymer membrane” as used herein is meant to denote a dry fibrillated membrane or a dry fibri Hated membrane assembly that has been through a wetting process.

[0087] The terms “non-dry”, “wet”, and “wetted” may be used interchangeably herein and are meant to refer to a dry fibrillated membrane or dry chromatography device before going through a drying process.

[0088] As used herein, the term “dry” may be defined as having a moisture content of less than or equal to 60% by mass of the dry fibrillated polymer membrane, or less than 55% by mass of the dry membrane, or less than 50% by mass of the dry membrane, or less than 45% by mass of the dry membrane, or less than 40% by mass of the dry membrane, or less than 35% by mass of the dry membrane, or less than 30% by mass of the dry membrane, or less than 25% by mass of the dry membrane, or less than 20% by mass of the dry membrane, or less than 15% by mass of the dry membrane, or less than 10% by mass of the dry membrane, or less than 5% by mass of the dry membrane, or less than 3% by mass of the dry membrane, or less than 1 % by mass of the dry membrane, or less than 0.1 % by mass of the dry membrane, or 0% by mass of the dry membrane.

[0089] As used herein, the term “dry” may also or alternatively be defined as having a moisture content less than or equal to 60% by mass of the dry affinity chromatography device, or less than 55% by mass of the dry device, or less than 50% by mass of the dry device, or less than 45% by mass of the dry device, or less than 40% by mass of dry device, or less than 35% by mass of the dry device, or less than 30% by mass of the dry device, or less than 25% by mass of the dry device, or less than 20% by mass of the dry device, or less than 15% by mass of the dry device, or less than 10% by mass of the dry device, or less than 5% by mass of the dry device, or less than 3% by mass of the dry device, or less than 1 % by mass of the dry device, or less than 0.1 % by mass of the dry device, or 0% by mass of the dry device.

[0090] As used herein, the term “about” denotes +/- 10% of the designated unit of measure.

[0091] The present disclosure is directed to dry fibrillated polymer membranes that have a moisture content less than or equal to 60% by mass of the dry membrane. The dry fibril lated polymer membranes may be positioned within affinity chromatography devices in a wound or a stacked configuration. The affinity chromatography device (containing the dry fibrillated polymer membrane(s)) may have a moisture content less than or equal to 60% by mass of the device. Additionally, the dry fibrillated polymer membrane contains inorganic particles (e.g., spherical or irregularly shaped) within the membrane and/or on the membrane. An affinity ligand may be bonded to the inorganic particles and/or to the dry fibrillated polymer membrane. In forming the dry fibrillated polymer membrane, a non-dry fibrillated polymer membrane may be dried through lyophilization. This process of drying through lyophilization enables a more simplified device manufacturing process, where chemical immobilization of ligand(s) onto the membrane can be done on the fibrillated membrane prior to integration into a chromatography device, thereby removing the in-situ chemical immobilization step from the device manufacturing process.

[0092] The affinity chromatography device and the dry fibrillated polymer membrane may be treated with ethylene oxide, gamma irradiation, x-ray irradiation, or any other treatment method that may result in the sterilization of the device or the membrane. In some embodiments, the dry fibrillated polymer membrane may be treated separately before assembly of the chromatography device. In some embodiments, the dry fibrillated polymer membrane may be treated with the chromatography device after assembly of the chromatography device.

[0093] The chromatography device (in any form, such as, but not limited to, affinity chromatography device, cassette, cassette assembly, manifold, manifold chromatography device, manifolded manifold, and/or manifolded manifold chromatography device, as well as any other chromatography device described herein) may have a moisture content less than or equal to 60% by mass of the dry affinity chromatography device, or less than about 55% by mass of the dry device, or less than about 50% by mass of the dry device, or less than about 45% by mass of the dry device, or less than about 40% by mass of the dry device, or less than about 35% by mass of the dry device, or less than about 30% by mass of the dry device, or less than about 25% by mass of the dry device, or less than about 20% by mass of the dry device, or less than about 15% by mass of the dry device, or less than about 10% by mass of the dry device, or less than about 5% by mass of the dry device, or less than about 3% by mass of the dry device, or less than about 1 % by mass of the dry device, or less than about 0.1 % by mass of the dry device, or less than about 0.01 % by mass of the dry device, or less than about 0.001 % by mass of the dry device, or 0% by mass of the device.

[0094] The dry fibrillated polymer membrane (in any form, such as, but not limited to, fibrillated polymer membrane, fibrillated membrane disc, stacked membrane assembly, wound membrane assembly, spiral wound membrane assembly, membrane assembly, and/or fibrillated membrane layers, as well as any other fibrillated polymer membrane described herein) may have a moisture content less than or equal to 60% by mass of the dry fibrillated polymer membrane, or less than about 55% by mass of the dry membrane, or less than about 50% by mass of the dry membrane, or less than about 45% by mass of the dry membrane, or less than about 40% by mass of the dry membrane, or less than about 35% by mass of the dry membrane, or less than about 30% by mass of the dry membrane, or less than about 25% by mass of the dry membrane, or less than about 20% by mass of the dry membrane, or less than about 15% by mass of the dry membrane, or less than about 10% by mass of the dry membrane, or less than about 5% by mass of the dry membrane, or less than about 3% by mass of the dry membrane, or less than about 1 % by mass of the dry membrane, or less than about 0.1 % by mass of the dry membrane, or less than about 0.01 % by mass of the dry membrane, or less than about 0.001 % by mass of the dry membrane, or 0% by mass of the dry membrane.

[0095] The dry fibrillated polymer membrane (in any form, such as, but not limited to, fibrillated polymer membrane, fibrillated membrane disc, stacked membrane assembly, wound membrane assembly, spiral wound membrane assembly, membrane assembly, and/or fibrillated membrane layers, as well as any other fibrillated polymer membrane described herein) may have a moisture content from about 0.001 % to 60% by mass of the dry fibrillated polymer membrane, from about 0.001 % to about 55% by mass of the dry membrane, or from about 0.01% to about 50% by mass of the dry membrane, or from about 0.05% to about 45% by mass of the dry membrane, or from about 0.1 % to about 40% by mass of the dry membrane, or from about 0.5 % to about 35% by mass of the dry membrane, or from about 0.6% to about 30% by mass of the dry membrane, or from about 0.7% to about 25% by mass of the dry membrane, or from about 0.8% to about 20% by mass of the dry membrane, or from about 0.9% to about 15% by mass of the dry membrane, or from about 1 % to about 10% by mass of the dry membrane. In some embodiments, the dry fibril lated polymer membrane has a moisture content of from about 0.001 % to about 5% by mass of the dry fibri Hated polymer membrane, or from about 0.01 % to about 4.5% by mass of the dry membrane, or from about 0.1 % to about 4% by mass of the dry membrane, or from about 1 .0% to about 3.5% by mass of the dry membrane, or from about 1.1 % to about 3% by mass of the dry membrane. In some embodiments, the dry fibril lated polymer membrane has a moisture content of from about 1 .3% to about 2% by mass of the dry membrane.

[0096] The dry chromatography device (in any form, such as, but not limited to affinity chromatography device, cassette, cassette assembly, manifold, manifold chromatography device, manifolded manifold, and/or manifolded manifold chromatography device, as well as any other chromatography device described herein) may have a moisture content from about 0.001% to 60% by mass of the dry affinity chromatography device, from about 0.001 % to about 55% by mass of the dry device, or from about 0.01 % to about 50% by mass of the dry device, or from about 0.05% to about 45% by mass of the dry device, or from about 0.1 % to about 40% by mass of the dry device, or from about 0.5 % to about 35% by mass of the dry device, or from about 0.6% to about 30% by mass of the dry device, or from about 0.7% to about 25% by mass of the dry device, or from about 0.8% to about 20% by mass of the dry device, or from about 0.9% to about 15% by mass of the dry device, or from about 1 % to about 10% by mass of the dry device. In some embodiments, the chromatography device has a moisture content from about 0.001 % to about 5% by mass of the dry chromatography device, or from about 0.01 % to about 4.5% by mass of the dry device, or from about 0.1 % to about 4% by mass of the dry device, or from about 1 .0% to about 3.5% by mass of the dry device, or from about 1.1 % to about 3% by mass of the dry device. In some embodiments, the chromatography device has a moisture content of from about 1 .3% to about 2% by mass of the dry device. [0097] In some embodiments, the chromatography devices have a dynamic binding capacity (DBC) greater than 35 mg/ml at 10% breakthrough at a residence time of 20 seconds. Further, the affinity chromatography devices may have a cycling durability of at least 100 cycles without exceeding an operating pressure of 0.3 MPa. In further embodiments, the affinity chromatography devices have an elution volume from 100 mAU to 100 mAU of about 1 to about 6 column volumes. In some embodiments, the dry affinity chromatography devices may improve the stability or viability of a target molecule being purified. According to some embodiments, the dry affinity chromatography devices may have a reduced storage risk as elimination of fluid inside the device reduces risk of microbial growth during storage. According to some embodiments, the dry affinity chromatography devices may have reduced shipping risk as elimination of fluid inside the device reduces risk in the event of an accidental rupture or leak due to a drop. In some embodiments, an affinity chromatography device containing a dry f ibril lated polymer membrane may also enable shorter lead times for shipping and distribution. After storage and/or shipping, the dry affinity chromatography devices can subsequently be readily wetted for use with bioprocessing fluids.

[0098] In some embodiments, dry affinity chromatography devices may have higher permeability and/or shelf-life stability at ambient conditions. The affinity chromatography devices according to the present disclosure may include other benefits such as uniform flow front in normal flow in multiple configurations (e.g., stacked, spiral wound, cassette, and parallel manifolds), enabling the user to incorporate a plurality of dry affinity chromatography devices configured in parallel manifold, and/or performance scalability by residence time. It is to be appreciated that the terms: “manifold” and “parallel manifold” are used interchangeably herein and are intended to define the same configuration. A dry affinity chromatography device containing a dry fibril lated polymer membrane therein enables a variety of treatment modes of the final device (e.g., capacity to be sterilized such as by ethylene oxide treatment, gamma treatment, x-ray treatment either before or after assembly). Such sterilization methods are more limited or not possible when working with incumbent wet article device forms.

[0099] In use, the dry affinity chromatography devices are first wetted using liquids appropriate for bioprocessing, including, but not limited to, aqueous buffer solutions, harvests, acids, bases, organic solvents, and other fluids that are commonly used and known to those of skill in the art in bioprocessing. It is to be appreciated that chromatography devices described herein are considered “dry” if the device contains a dry fibrillated polymer membrane in any form.

[00100] FIGS. 1 and 2 are an exploded view and a cross-sectional view, respectively, of a chromatography device 100 containing a wound membrane according to at least one embodiment. In forming the chromatography device 100, in some embodiments, at least one inner intermediate material 200 may be circumferentially positioned against (e.g., wound around) a cylindrical core 150 to a desired width or to a pre-designated amount. A dry fibrillated polymer membrane containing therein spherical and/or non-spherical inorganic particles 210 is then wound around the core 150 over the inner intermediate material 200 to a desired width or to a pre-designated amount. An outer layer of at least one outer intermediate material 220 is then circumferentially positioned on (e.g., wound around) the dry fibrillated polymer membrane 210 to a desired width or to a pre-designated amount. Herein, the combination of the inner intermediate material 200, the dry fibrillated polymer membrane 210, and the outer intermediate material 220 will be referred to as the “wound membrane assembly”. In some embodiments, the “wound membrane assembly” may also contain any combination of polymer(s) and/or polymer intermediate material(s) wrapped around a cylindrical core. The cylindrical core 150 may have a hollow or solid interior. In either instance, the core 150 contains a solid outer wall so that an aqueous mixture flowing through the chromatography device 100 flows within an inner flow channel 140 formed of the inner intermediate material 200 and not into the core 150. The use of a hollow core 150 reduces the amount of material used to form the core 150, reduces the weight of the device 100, and reduces manufacturing costs.

[00101] As depicted in FIG. 1 , the wound membrane assembly 110 includes the inner intermediate material(s) 200, the dry fibrillated polymer membrane 210, and the outer intermediate matehal(s) 220. The wound membrane assembly 110 and central core 150 may be positioned within a housing 50. In some embodiments, the housing 50 is cylindrical. As depicted in FIG. 2, the outer intermediate material(s) 220 forms an outer flow channel 130 and the inner intermediate material(s) 200 form an inner flow channel 140. It is to be appreciated that the intermediate material(s) 200, 220 in the embodiments described herein may be different or they may be the same. Additionally, two or more intermediate materials may be used to form one or both of the outer flow channel 130 and the inner flow channel 140. Non-limiting examples of suitable materials forming the housing for the chromatography devices described herein, include, but are not limited to, polyurethane, stainless steel, polypropylene, acrylonitrile butadiene styrene (ABS), polyethylene terephthalate (PET), polyether ether ketone (PEEK), cyclic olefin copolymer (COC), and polyethylene terephthalate glycol (PETG). Additionally, the shape of the housing unit is not limiting, and may take any form so long as the form encapsulates the affinity chromatography devices and/or the cassette(s) (described below) and/or the manifold(s) (described below).

[00102] In use, the dry fibrillated polymer membrane 210 is first wetted as described above. After wetting, an aqueous mixture flows into the inlet 80 positioned within inlet cap 60. The mixture flows over the distributor cap 65 and is directed towards the outer flow channel 130 formed by the outer intermediate material(s) 220. The distributor cap 65 directs the aqueous mixture roughly 90 degrees from the feed direction towards the outer flow channel 130 (/.e., intermediate material(s) 220). This redirection promotes a more uniform flow of the aqueous mixture into the outer flow channel 130. The outer intermediate material(s) 220 forming the outer flow channel 130 is located between the housing 50 and the wound dry fibrillated polymer membrane 210. The distributor cap 65 may be a polyolefin or be coated with a polyolefin.

[00103] The aqueous mixture flows through the outer flow channel 130 (/.e., outer intermediate material(s) 220) and across the wound “wetted” fibrillated polymer membrane 210 in a normal direction (e.g., from the outer flow channel 130 to the inner flow channel 140). As the aqueous mixture is passed in a normal flow from the outer flow channel 130 (/.e., outer intermediate material(s) 220) and across the wound wet fibrillated polymer membrane 210, the affinity ligand reversibly binds to the targeted molecule, thereby effectively removing it from the aqueous mixture. The aqueous mixture minus the targeted molecule then enters the inner flow channel 140 (/.e., inner intermediate material(s) 200) located between the solid outer wall of the central core 150 and the wound wet fibrillated polymer membrane 210. The aqueous mixture then is redirected at the bottom of the inner flow channel 140 by an outlet cap 75. The aqueous mixture then flows out of the chromatography device 100 through outlet 85 located within the outlet cap 75.

[00104] The diameter and/or height of the central core 150 and/or the width and/or height of the dry fibrillated polymer membrane and/or intermediate material(s) can be adjusted to achieve a larger volume without negatively impacting performance of the chromatography device. Additionally, the targeted molecule may be removed from the affinity ligand, for example, by passing a fluid that has a lower pH through the chromatography device, as is known by those of ordinary skill in the art.

[00105] The intermediate material(s) 200 and 220 are not particularly limiting so long as the aqueous mixture is able to flow therethrough. Some non-limiting examples of suitable intermediate materials include, but are not limited to, a porous fluoropolymer film or a porous non-fluoropolymer film (e.g., a porous polypropylene or other porous polyolefin film), a porous non-woven material, or a porous woven material. In some embodiments, the intermediate film is a thermoplastic or thermoset polymer film. In some embodiments, the wound membrane assembly includes an integrated inlet end cap 60 and/or an integrated outlet end cap 75 at opposing ends of the housing 50 to form an integrated, reusable chromatography device. In some embodiments, the inlet or inlet end cap 60 and outlet or outlet end cap 75 may be positioned on the same side of the dry chromatography device 100.

[00106] In some embodiments, the affinity chromatography device 100 or the dry fibrillated polymer membrane 210 may be treated with ethylene oxide, gamma irradiation, x-ray irradiation, or any other sterilization/treatment method that results in the sterilization of the device 100 or membrane 210. In some embodiments, the dry fibrillated polymer membrane 210 may be sterilized separately before assembly of the affinity chromatography device 100. In some embodiments, the dry fibrillated polymer membrane 210 may be sterilized after assembly of the affinity chromatography device 100.

[00107] In some embodiments, such as is depicted generally in FIG. 3, the chromatography device 300 contains a dry fibrillated polymer membrane configured as individual dry fibrillated polymer membrane discs 340 stacked upon each other to form a stacked membrane assembly 320. The dry fibrillated polymer membrane discs 340 may be positioned in a stacked configuration by simply laying the dry fibrillated polymer membrane discs 340 on top of each other. Alternatively, the dry fibrillated polymer membrane discs 340 may be stacked and subsequently laminated together with heat and/or pressure or by any other conventional methods. The stacked membrane assembly 320 described herein is with respect to dry fibrillated polymer membrane discs for ease of explanation. The dry fibrillated polymer membrane may be formed in other geometric shape(s) and/or non-geometric shape(s) and are considered to be within the purview of this disclosure. It is to be appreciated that the chromatography device 300 is considered “dry” if the affinity chromatography device 300 contains a dry fibrillated polymer membrane or dry fibrillated polymer membrane discs 340.

[00108] Continuing with FIG. 3, the affinity chromatography device 300 includes at least one upper intermediate material(s) 360 positioned at the inlet side of the stacked membrane assembly 320 and at least one lower intermediate material(s) 380 positioned at the outlet side of the stacked membrane assembly 320. The upper and lower intermediate material(s) 360, 380, respectively, may be the same or different. Similar to the wound membrane assembly discussed above, the intermediate material(s) 360, 380 used to form the stacked membrane assembly 320 is not particularly limiting so long as the aqueous mixture is able to flow therethrough. Non-limiting examples of suitable intermediate materials include, but are not limited to, a porous fluoropolymer film or a porous non-fluoropolymer film (e.g., a porous polypropylene or other porous polyolefin film), a porous non-woven material, or a porous woven material.

[00109] The stacked membrane assembly 320 may be disposed within a housing 350 having an inlet cap 365 and an outlet cap 375 disposed at opposing ends of the housing 350. In some embodiments, for example, where the dry fibrillated polymer membrane has the form of a disc or circular shape, the housing 350 is cylindrical, although any geometry that is capable of housing the stacked membrane assembly and achieving a desired dynamic binding capacity is considered to be within the purview of this disclosure. In some embodiments, the intermediate material(s) 360, 380, the housing 350, the inlet cap 365, and the outlet cap 375 may be formed of a thermoplastic polymer such as polypropylene, polyethylene, or other polyolefin(s). Alternatively, one or both of the intermediate material(s) 360, 380 may be formed of an inorganic or metallic material, so long as the porous intermediate material(s) 360, 380 do not hinder the operation of the chromatography device.

[00110] The dry fibrillated polymer membrane discs 340 in the stacked membrane assembly 320 may be adhered to the housing 350 at the inner walls of the housing 350 via any conventional process (e.g., melt sealing or use of a sealant) that prevents flow between the periphery of the dry fibrillated polymer membrane discs 340 and the housing 350. The inlet cap 365 and the outlet cap 375 may be sealed to the housing 350 by a similar or identical process. The inlet cap and the outlet cap 365, 375 includes an inlet 380 and an outlet 385, respectively, to permit the flow of an aqueous mixture into and out of the affinity chromatography device 300. Specifically, the inlet 380 permits fluid flow into the housing 350 and the outlet 385 permits fluid flow out of the housing 350. The dry fibrillated polymer membrane discs 340 remains dry prior to use of the chromatography device 300. Prior to use, the dry affinity chromatography device 300 or dry fibrillated polymer membrane/dry fibrillated membrane discs 340 are wetted using liquids appropriate for bioprocessing, including, but not limited to, aqueous buffer solutions, harvests, acids, bases, organic solvents, and other fluids that are commonly used and known to those of skill in the art in bioprocessing.

[00111] In use, the aqueous mixture flows sequentially through the upper intermediate material(s) 360, through the “wetted” fibrillated polymer membrane discs 340 forming the stacked membrane assembly 320, and through the lower intermediate material(s) 380. As the aqueous mixture is passed through the “wetted” fibrillated polymer membrane discs 340, the affinity ligand reversibly binds to the targeted molecule, thereby effectively removing it from the aqueous mixture. The targeted molecule may be removed from the affinity ligand, for example, by passing a fluid that has a lower pH through the device, as is known by those of skill in the art.

[00112] In some embodiments, the affinity chromatography device 300 and/or the dry fibrillated polymer membrane discs 340 may be treated with ethylene oxide, gamma irradiation, x-ray irradiation, or other treatment methods that may result in sterilization of the device 300 (including sterilization of the dry fibrillated membrane discs 340). In some embodiments, the stacked membrane assembly 320 or the dry fibrillated polymer discs may be sterilized with any of the forementioned sterilization methods prior to assembling the affinity chromatography device.

[00113] The total number of dry fibrillated polymer membranes present in a stacked membrane assembly is not particularly limited and depends on the desired end use and/or desired mass transit flow within the stacked membrane assembly. The 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) total dry fibrillated polymer membranes. It is to be appreciated that hundreds or even thousands of dry fibrillated polymer membranes may be present in the stacked membrane assembly.

[00114] As mentioned previously, the dry fibrillated polymer membrane(s) contain(s) therein inorganic particles. In some embodiments, the dry fibrillated polymer membrane may contain one or more than one type of inorganic particle and/or one or more than one nominal particle size within the dry fibrillated polymer membrane. Nonlimiting examples of suitable inorganic particles include, but are not limited to, silica, zeolites, hydroxyapatite, metal oxides, and combinations thereof. It is to be understood that the term “silica” as used herein is meant to describe a silicon dioxide that does not contain any measurable amount of boron or contains no boron as measured by x-ray photoelectron spectroscopy (XPS). Additionally, the inorganic particles may be either solid or porous and may have a variety of sizes and shapes.

[00115] The inorganic particles may have a nominal particle size from about 0.1 microns, about 0.5 microns, from about 1 micron, from about 5 microns, about 10 microns, from about 15 microns, from about 20 microns, or from about 25 microns or more. Further, the inorganic particles may be monodisperse or polydisperse.

[00116] The dry fibrillated polymer membrane(s) in both the wound membrane assembly and the stacked membrane assembly may contain therein spherical inorganic particles, or particles having a spherical configuration. As used herein, the term “spherical” is meant to denote that the inorganic particle has a round or nearly round shape where the distance from the center of the inorganic particle to the outer edge of the particle at any point is the same or nearly the same distance. In some embodiments, the spherical inorganic particles have a particle size distribution that has a 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 spherical inorganic particles have a nominal particle size that may be about 5 microns, about 10 microns, about 15 microns, about 20 microns, and combinations and blends thereof. In some embodiments, the spherical inorganic particles are polydisperse.

[00117] In some embodiments, the affinity ligand is covalently bonded to the inorganic particles. In another embodiments, the affinity ligand is covalently bonded to the dry fibrillated polymer membrane. In a further embodiment, the affinity ligand may be bound to both the dry fibrillated polymer membrane and the inorganic particle(s). The affinity ligand may be a protein, antibody, or polysaccharide that reversibly binds to a targeted protein or antibody. In one embodiment, the affinity ligand is a protein that reversibly binds, for example, to an 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, Protein L, or a polysaccharide that reversibly binds to a protein or a protein fragment to which it is specific. Affinity ligands for use in the affinity chromatography device may include, but are not limited to, Protein A, Protein G, Protein L, human Fc receptor protein, antibodies that specifically bind to other proteins, and heparin. The affinity ligand may be native, recombinant, or synthetic. In some embodiments, the affinity ligand may be an oligosaccharide or an oligonucleotide that reversibly binds to nucleic acids including, but not limited to, mRNA. In some embodiments, the affinity ligand may be an antibody or a polysaccharide that reversibly binds to viral vectors including, but not limited to, adeno-associated viruses. In yet another embodiment, the affinity ligand is a metal affinity ligand that reversibly binds to His-Tagged Proteins.

[00118] According to some embodiments, the dry fibrillated polymer membrane includes an expanded polytetrafluoroethylene membrane, an expanded modified polytetrafluoroethylene membrane, an expanded tetrafluoroethylene copolymer membrane, or an expanded polyethylene membrane. In some embodiments, the dry fibrillated polymer membrane includes an expanded polytetrafluoroethylene membrane or an expanded polyethylene membrane.

[00119] Although embodiments of the wound membrane assembly 110 and stacked membrane assembly 320 are described herein, it is to be appreciated that any number of dry fibrillated polymer membranes as well as any and all combinations of types of dry fibrillated polymer membranes, types of spherical inorganic particles, sizes of spherical inorganic particles, and orientations of the dry fibrillated polymer membrane(s) within the membrane assemblies 110, 320 are within the scope of this disclosure. Also, some or all of the dry fibrillated polymer membranes may vary in composition, thickness, permeability, etc. from each other.

[00120] FIG. 4A is a cross-sectional view of a chromatography device 400 containing a cassette assembly 401 ; FIG. 4B is an exploded view of the chromatography device 400 shown in FIG. 4A. The cassette assembly 401 includes a dry fibrillated polymer membrane assembly containing therein inorganic particles. The dry fibrillated polymer membrane assembly may be configured as individual dry fibrillated polymer membrane layers 440 stacked upon, or otherwise parallel to, each other to form a stacked membrane assembly 410. In some embodiments, the dry fibrillated polymer membrane layers 440 may be positioned in a stacked configuration by simply laying the dry fibrillated polymer membrane layers 440 adjacent to each other. Alternatively, the dry fibrillated polymer membrane layers 440 may be stacked and subsequently laminated together (e.g., intermittently spaced across the membrane layers or joined at the edges of each membrane layer). As shown, the affinity chromatography device 400 (e.g., cassette assembly 401 ) includes a fluid inlet channel 402 connected to an inlet housing 404 and a fluid outlet channel 406 connected to an outlet housing 408. The fluid inlet channel 402 is fluidly connected to the fluid outlet channel 406. In some embodiments, the device 400 includes a dry fibrillated polymer membrane assembly 410 containing therein at least one inorganic particle positioned between the inlet housing 404 and the outlet housing 408. It is to be appreciated that the chromatography device 400 is considered “dry” if the device contains a dry fibrillated polymer membrane.

[00121] The chromatography device 400 optionally includes at least one inlet side intermediate material 460 positioned at the inlet side of the dry stacked membrane assembly 410 and at least one outlet side intermediate material 480 positioned at the outlet side of the dry stacked membrane assembly 410. The inlet side and outlet side intermediate materials 460, 480, respectively, may be the same or different. Similar to the wound and stacked membrane assemblies discussed above, the inlet side and outlet side intermediate materials 460, 480 used to form the cassette stacked membrane assembly 410 are not particularly limiting so long as the aqueous mixture is able to flow therethrough. Non-limiting examples of suitable intermediate materials include, but are not limited to, a porous fluoropolymer film or a porous non- fluoropolymer film (e.g., a porous polypropylene or other porous polyolefin film), a porous non-woven material, or a porous woven material. In some embodiments (not shown), the inlet side and outlet side intermediate materials 460, 480 are not present. [00122] In some embodiments, at least one of the dry fibrillated polymer membrane layers 440 and the inorganic particles have covalently bonded thereto an affinity ligand that reversibly binds to a targeted molecule. In some embodiments, the affinity chromatography device 400 may be treated with ethylene oxide, gamma irradiation, x-ray irradiation, or other treatment methods known to those of skill in the art that sterilize the dry fibrillated polymer membrane layers 440 or the chromatography device 400.

[00123] As shown in FIGS. 4A and 4B, the inlet housing 404 and the outlet housing 408 are substantially planer and stacked on, or otherwise parallel to, opposing sides of the dry stacked membrane assembly 410, thereby forming the cassette assembly 401. The dry stacked membrane assembly 410 contains the dry fibrillated polymer membrane layers 440. In some embodiments, the inlet housing 404 and the outlet housing 408 have substantially the same shape as the dry fibrillated polymer membrane assembly 410. For example, as shown in FIG. 4A and FIG. 4B, both housings 404, 408 and the dry stacked membrane assembly 410 are of a substantially rectangular shape. In some embodiments, the housings 404, 408 and the dry stacked membrane assembly 410 may be of any shape so long as the housings 404 and 408 fully encompass the dry stacked membrane assembly 410.

[00124] In some embodiments, the dry stacked membrane assembly 440 (and the dry fibrillated polymer membrane layers 440) may include a perimeter sealing material 412 sealing the edges of the stacked membrane assembly 410. The perimeter sealing material 412 is not particularly limiting so long as the aqueous mixture is able to flow through the stacked membrane assembly 410. Some non-limiting examples of suitable perimeter sealing materials include, but are not limited to, polyolefins, thermoplastic elastomers, or combinations thereof.

[00125] In some embodiments, for example, and as shown in FIG. 4B, the inlet side and outlet side intermediate materials 460, 480 may be outside of the perimeter sealing material 412. In some embodiments, the inlet side and outlet side intermediate materials 460, 480 may each include a separate intermediate perimeter sealing material different from the perimeter sealing material 412 sealing the edges of the stacked membrane assembly 410. In yet some embodiments (not shown), the inlet side and outlet side intermediate materials 460, 480 may be inside, and thus sealed within, the perimeter sealing material 412.

[00126] FIG. 5 is a side view of a parallel manifold chromatography device 500 having a plurality of chromatography devices (each of which is a cassette assembly), where each chromatography device includes a dry fibril lated polymer membrane containing therein inorganic particles. In some embodiments, the plurality of chromatography devices may include a mix of “wetted” and dry fibri Hated polymer membranes.

[00127] As shown, the manifold chromatography device 500 includes 6 cassette assemblies 501 a-f (e.g., the cassette assembly 401 in FIG. 4A) arranged in a parallel configuration. In some embodiments, a manifold may include anywhere between 2 to 20 cassette assemblies parallel to each other. In some embodiments, a manifold may include more than 20 cassette assemblies parallel to each other, so long as there is a similar distribution of flow and permeability across the cassette assemblies. This similarity across the cassette assemblies allows for scalability of device volume and performance.

[00128] In some embodiments, the fluid inlet channel 502 may include an optional flow distributer 520 disposed upstream of a first device 501 a of the plurality of devices 501 a-f. The optional flow distributor 520 may enable more uniform flow and help facilitate flow distribution though the fluid inlet channel 502.

[00129] In some embodiments, the manifold chromatography device 500 may include an optional gas separation device 530 (e.g., an integrated air trap) upstream of the fluid inlet channel 502. It is to be appreciated that the fluid inlet channel 502 and the fluid outlet channel 504 may be on opposite sides of the manifold of chromatography devices, the same side, or in other configurations, so long as fluid flow through the individual chromatography devices 501 a-f occurs in parallel.

[00130] FIG. 6 is a cross-sectional view of a manifolded manifold 650 having at least two manifolds 600A and 600B (e.g., the manifold of chromatography devices 500 in FIG. 5) positioned within an optional housing 620. Each manifold contains cassette assemblies that each include a dry fibrillated polymer membrane containing therein inorganic particles.

[00131] As shown in FIG. 6, the manifolded manifold 650 includes 2 manifolds 600A and 600B (e.g., the manifold of chromatography devices 500 in FIG. 5) arranged in a parallel configuration. In some embodiments, a manifolded manifold may include anywhere between 2 to 20 manifolds parallel to each other. In some embodiments (not shown), a manifolded manifold may include more than 20 manifolds parallel to each other. It is to be appreciated that the fluid inlet channel 652 and the fluid outlet channel 654 may be on opposite sides of the manifold of chromatography devices, the same side, or in other configurations, so long as fluid flow through the individual manifolds 600A and 600B occurs in parallel. In some embodiments, the manifolds 600A, 600B may each include a fluid outlet channel 654A, 654B, and an optional gas separation device 630A, 630B (e.g., an integrated air trap) upstream of the fluid inlet channels 652A, 652B. In further embodiments (not shown), an optional gas separation device may be disposed upstream of the manifolded manifold device fluid inlet channel 652. [00132] FIG. 7 is a side, cross sectional view of a manifold 750 having a plurality of chromatography devices, each of which contains a wound membrane assembly. Each wound membrane assembly includes a dry fibrillated polymer membrane containing therein inorganic particles. The chromatography devices are arranged in a parallel configuration. The chromatography devices described with respect to FIG. 7 are considered to be dry chromatography devices if they include a dry fibrillated polymer membrane or a dry wound membrane assembly.

[00133] In some embodiments, the affinity chromatography devices described herein may be utilized in a manifold 750 with wound membrane affinity chromatography devices 700, 701 (e.g., the chromatography device 100 set forth in FIGS. 1 and 2) arranged in a parallel configuration, such as is generally depicted in FIG. 7. In some embodiments, the manifold 750 may include an optional gas separation device 730 (e.g., an integrated air trap) upstream of the fluid inlet 740. In some embodiments, the manifold 750 is positioned within a housing 720.

[00134] In use, the dry stacked membrane assembly or dry fibrillated polymer membranes positioned in each of the chromatography device is first wetted using liquids appropriate for bioprocessing, including, but not limited to, aqueous buffer solutions, harvests, acids, bases, organic solvents, and other fluids that are commonly used and known to those of skill in the art in bioprocessing. Next, an aqueous mixture flows into the fluid inlet 740. The fluid inlet 740 divides the flow of the aqueous mixture into at least two inlet tubes 760, 761. The split aqueous mixture in inlet tubes 760, 761 then flows into chromatography devices 700, 701 , where a targeted molecule is captured by the affinity ligand positioned on a fibrillated polymer membrane and/or inorganic particle. An aqueous solution, which is the aqueous mixture minus the targeted molecule captured by the affinity ligand, flows out of the chromatography devices 700, 701 through outlet tubes 780, 781 , respectively. The aqueous solution in outlet tubes 780, 781 are combined in distribution element 790 and reconstituted into a single aqueous solution. [00135] It is to be appreciated that two chromatography devices 700, 701 are shown in FIG. 7 for illustration only and a plurality, more than two, chromatography devices described herein may be utilized in a manifold in a parallel configuration so long as there is a similar distribution of flow and permeability across the chromatography devices. This similarity across the affinity chromatography devices allows for scalability of device volume and performance. In addition, the affinity chromatography devices may be dropped into a parallel configuration system without any changes or additions to the manifold 750. The fluid inlet 740 and the fluid outlet 790 may be on opposite sides of the manifold of chromatography devices, the same side, or in other configurations, so long as fluid flow through the individual chromatography devices 700, 701 occurs in parallel. [00136] FIG. 8 is a cross-sectional view of two manifolds containing a wound dry fibrillated polymer membrane assembly in a parallel configuration in accordance with at least one embodiment. As shown in FIG. 8, a manifolded manifold 26 contains two manifolds 6a, 6b in a parallel configuration. The manifolds 6a and 6b (e.g., the manifold 750 in FIG. 7) each contain at least two affinity chromatography devices, 26a, 26b, and 26c, 26d. The ability to utilize at least two manifolds in parallel advantageously allows for an increase in volume capacity while utilizing the chromatography devices described herein. In some embodiments, the manifolded manifold 26 may include an optional gas separation device 830 (e.g., an integrated air trap) upstream of the fluid inlet 30.

[00137] Turning to FIGS. 10A and 10B, a dry fibrillated membrane in the form of a stacked membrane assembly as described above, may be affixed to a multi-well plate 1000 containing a porous surface 1030 separating a lower chamber that can be operated at reduced pressure from an upper chamber that is operated at a higher (e.g., atmospheric) pressure. In the embodiment depicted in FIG. 10A, a stacked membrane assembly formed of dry fibrillated polymer membranes 1020, each of which contains inorganic particles may be utilized.

TEST METHODS

Method for Determining Moisture Content of Membranes

[00138] Membrane moisture content was determined following the guidelines defined by ASTM Standard E1868-10 Standard Test Methods for Loss-On-Drying for Thermogravimetry. A Mettler Toledo HB43 Moisture Analyzer (Mettler-Toledo GmbH, CH-8606 Greifensee, Switzerland) was used for all data collection. A consistent membrane area of 109.5 cm² was used, die cut as six 2 cm diameter circles. For wet samples, each circle was briefly placed on a low-lint wipe for a few seconds to remove surface moisture before being loaded into the instrument for measurement. Each sample was heated at 110 °C for a total of 2 hours. Values were reported as Loss-On- Drying (LOD) in units of % by mass.

Method for Air Permeability, Thickness, and Density of Membranes

[00139] To determine the density and thickness of each membrane sample, a 1- inch diameter (5.3 cm²) circle was punched from each membrane. This sample was weighed and then the thickness was measured with a Mitutoyo Litematic VL-50 thickness gauge (Mitutoyo America Corporation, Aurora, IL). The membrane was characterized for air permeability using a Gurley Model 4340 Automatic Densometer (Gurley Precision Instruments, 514 Fulton Street, Troy, NY 12180). Density was calculated by weighing each die cut circle and dividing by the sample volume, calculated from the die cut diameter and the measured thickness.

Method for Dynamic Bind and Elute Behavior Determination for Membranes [00140] To determine the dynamic binding capacity of each membrane sample, two 25mm diameter (4.9 cm²) circles were die cut from each membrane. These samples were stacked and held in a filter clamp to allow for controlled flow of solutions through the membrane.

[00141] Two 25 mm disks of the membrane sample were positioned between a support screen (VWR® Support Screen, Stainless Steel, 89428-948) and a glass funnel (VWR® Glass Funnel, 15 ml, 89428-938), in a 125 ml Graduated Filter Flask assembly (VWR® Filter Flask Graduated, 125 ml, 89428-978).

[00142] A sample vial (VWR® Sample Vial, Clear Borosilicate Glass, Screw- thread, 20 ml, 66009-567) was placed into the 125ml filter flask (VWR® Graduated Filter Flask, 125 ml, 89428-978) with the outlet of the glass base (VWR® Glass Base for Stainless Steel Support, 89428-942) positioned inside the sample vial. The support screen (VWR® Support Screen, Stainless Steel, 89428-948) was positioned on the glass base. The membrane sample was positioned between the support screen and the glass funnel (VWR® Glass Funnel, 15 ml, 89428-938) and held by the aluminum clamp (VWR® Aluminum Clamp, 89428-944).

[00143] A single bind and elute cycle to determine the dynamic bind and elute performance of the membrane was performed using the following protocol. Solutions described in Table A were applied to the glass funnel in the volumes and method order described in Table B. The eluate from each method step was collected in a dedicated sample vial (VWR® Sample Vial, Clear Borosilicate Glass, Screw-thread, 20 ml, 66009- 567) as indicated in Table B. The eluate sample vial was replaced after each method step. Each solution flowed through the membrane under the force of gravity.

Table A

Table B

[00144] Concentration of the mAb in each elution sample was calculated by measuring the absorption at a wavelength of 280 nm using a Hitachi U-2900 spectrophotometer (Hitachi High-Tech in America, 10 North Martingale Road, Suite 500, Schaumburg, Illinois 60173-2295).

[00145] Concentration of the mAb in the Target Analyte Feed and in the flowthrough and wash samples was calculated by measuring the absorption at a wavelength of 280 nm using a Hitachi U-2900 spectrophotometer (Hitachi High-Tech in America, 10 North Martingale Road, Suite 500, Schaumburg, Illinois 60173-2295) [00146] Beer’s law (Eq-01 ) was then used to calculate the mAb concentration of the samples.

Eq-01 A = e x L x C where:

A = absorbance e = 1.47 mL g^-1 cm^-1 (extinction coefficient of target analyte, trastuzumab biosimilar)

L = sample path length

C = solution concentration

Method for Lyophilization of Membranes

[00147] To dry each Protein A immobilized membrane, the membranes were suspended in place and thoroughly rinsed with deionized (DI) water to ensure most of the solvent to be evaporated was water. A final soak in water was completed and each membrane was subsequently frozen for 24 hours at -40 °C in a Revco DXF-40040A ultra-low temperature freezer (Thermo Fisher Scientific, 81 Wyman Street, Waltham, MA 02451 ). Each suspended frozen membrane was transferred into a VirTis 25L Genesis SQ Super ES 55 lyophilizer (SP Industries, 935 Mearns Road, Warminster, PA 18974), where both the condenser and shelves were frozen prior to beginning the cycle, then the steps outlined in Table C were followed to dry the membranes after initial shelf (< - 35°C) and vacuum (< 100mT) conditions were achieved per the programmed settings of the cycle.

Table C

[00148] After completing the lyophilization cycle, each dry membrane was characterized for moisture content according to the Method for Determining Moisture Content of Membranes.

Method for Ethylene Oxide (EO) Treatment of Membranes

[00149] Each dry membrane that underwent an ethylene oxide (EO) treatment was sectioned and placed into sterilization pouches (Part Number PG-7514, plastcareusa, Canoga Park, CA) after being dried following the Method for lyophilization of membranes.

[00150] A 3M Steri-Vac™ Sterilizer GS8X-1 D (3M Health Care, St. Paul, MN) was used to perform an EO treatment for some membranes, with treatment conditions representative of an EO sterilization cycle. The EO treatment cycle injected 170 grams of EO into the sterilizer containing the membranes using a Steri-Gas™ EO Gas Cartridge (Part Number 8-170, 3M Health Care, St. Paul, MN) and the sterilizer was held at a temperature of 40°C for 3 hours. The details of this cycle can be observed in Table D.

Table D

Method for Determining the Dynamic Binding Capacity at 10% Breakthrough for Device Articles

[00151] The chromatography device was inserted in the flow path of an AKTA™ Pure (Cytiva, Marlborough, MA) liquid chromatography system and a single cycle consisting of the following protocol was performed. Table E sets forth the solutions utilized, and Table F sets forth the protocol steps to determine the dynamic binding capacity at 10% breakthrough.

Table E

*Clean-in-Place

Table F

Method for Determining Liquid Permeability for Device Articles

[00152] The liquid permeability of the chromatography devices was determined using Darcy’s law. Individual devices were characterized for bed cross sectional area and bed length. Solution A was used as the liquid and was characterized for viscosity The pressure drop across the column as a function of liquid flux was measured on an AKTA™ Pure liquid chromatography system (Cytiva, Marlborough, MA). Method for Initial Wetting of Dry Device Articles

[00153] Dry device articles were wetted prior to characterization tests. A chromatography device was inserted into the flow path of an AKTA Pure liquid chromatography system, and a 50mM sodium phosphate 150mM sodium chloride pH 7.4 buffer was fed through the device, first in an up-flow direction followed by a downflow direction, following the flow rate and volumes in Table G below.

Table G

Method for determining moisture content of Device Articles

[00154] Dry device articles were weighed following assembly to obtain device dry mass. The device articles were then weighed again after initial wetting and permeability and DBC data was collected. The device wet mass represents the device when filled with the storage solution specified in Table E.

Method for Ethylene Oxide (EO) Treatment of Device Articles

[00155] Each dry device article that underwent an ethylene oxide (EO) sterilization treatment was placed into sterilization pouches (Part Number PG-7514, plastcareusa, Canoga Park, CA).

[00156] A 3M Steri-Vac™ Sterilizer GS8X-1 D (3M Health Care, St. Paul, MN) was used to perform an EO treatment for some devices, with treatment conditions representative of an EO sterilization cycle. The EO treatment cycle injected 170 grams of EO into the sterilizer containing the devices using a Steri-Gas™ EO Gas Cartridge (Part Number 8-170, 3M Health Care, St. Paul, MN) and the sterilizer was held at a temperature of 40°C for 3 hours. The details of this cycle can be observed in Table H.

Table H

Method for Determining Elution Volume from 100m AU to 100m AU for Device Articles

[00157] The wetted chromatography device was inserted in the flow path of an AKTA™ Pilot 600 (Cytiva, Marlborough, MA) liquid chromatography system and a single cycle consisting of the following protocol was performed. Table I sets forth the solutions utilized, and Table J sets forth the purification protocol steps utilized. The resulting UV absorbance at 280nm chromatogram, generated by following the purification protocol in Table J, was used to determine the elution volume from l OOmAU to 100mAU.

Table I

*C ean-in-Place

Table J

[00158] The resulting chromatogram of UV absorbance at 280 nm plotted as a function of cumulative volume passed through the device, generated by following the purification protocol in Table J, was used to determine the 100 mAU to 100 rnALI elution volume of the device article, using equation 02 (Eq-02).

[00159] Eq-02 £^ioo to ioo ⁼ CV_{10Q end} — CV_{10Q start} where:

EV100 to 100 = Elution Volume from 100 mAU to 100 mAU,

CVwo start = Cumulative volume of buffer solution and CHO Cell Harvest (in column volumes, CV) that has passed through the device by the start of the elution, defined as the point at which UV absorbance at 280nm rises above 100 mAU when feeding elution buffer to the device per Table J, step 4, and

CV100 end = Cumulative volume of buffer solution and CHO Cell Harvest (in column volumes, CV) that has passed through the device by the end of the elution, defined as the point at which UV absorbance at 280nm drops below 100 mAU when feeding elution buffer to the device per Table J, step 4.

EXAMPLES

[00160] Membrane A was a porous polytetrafluoroethylene (ePTFE) membrane having 15 mass percent ePTFE and 85 mass percent porous spherical silica particles. Membrane B was a porous ePTFE membrane having 15 mass percent PTFE and 85 mass percent porous irregularly shaped silica particles. Membranes A and B were the same as those described in U.S. Patent Publication No. 2023/0356109 to Clinger, et al. Table K lists some of the physical characteristics of the two porous ePTFE membranes

Table K

[00161] The membranes were then treated in a manner to covalently bond (immobilize) Protein A to the ePTFE membranes. This manner is representative of that typical to those skilled in the art and is further described in U.S. Patent No. 10,525,376 to McManaway, et al. and in U.S. Patent No. 10,526,367 to McManaway, et al, generating Protein A immobilized membranes C and D, respectively. [00162] Protein A immobilized membranes were dried via lyophilization as set forth in the Method for Lyophilization of Membranes described above, generating dry Protein A immobilized membranes E and F, respectively.

[00163] Sections of dry immobilized membranes E and F underwent an ethylene oxide treatment as set forth in the Method for Ethylene Oxide (EO) Treatment of membranes described above, generating dry Protein A immobilized ethylene oxide treated membranes G and H, respectively.

[00164] The membranes whose manufacture was described above were tested to evaluate their moisture content, density, thickness, and dynamic bind and elute behavior using the protocols described in the Test Methods set forth herein. For each dry membrane, the air permeability was also measured. The performance of each of these affinity chromatography membranes is shown in Table L.

Table L

[00165] Non-dry membranes C and D whose manufacture was described above were used to manufacture non-dry reference affinity chromatography devices in stacked membrane and parallel manifolded formats. Dry membranes E through H whose manufacture was described above were used to manufacture dry affinity chromatography devices in stacked membrane, spiral-wound, and parallel manifolded formats. Membranes and conditions associated with each non-dry and dry affinity chromatography device are summarized in Table M. The dry affinity chromatography devices were wetted using the method described above with a 50m L sodium phosphate with 150mM sodium chloride buffer solution then tested to evaluate their liquid permeability and twenty (20) second residence time dynamic binding capacities using the protocols described in the Test Methods. The performance of these devices is reported in Table N.

Table M

Table N

[00166] Device T was further characterized for elution volume performance using the Method for Determining Elution Volume from 100 mAU to 100 rnALI for Device Articles. The performance of Device T is shown in Table O. The UV absorbance at 280nm chromatogram associated with the characterization of Device T using the Method for Determining Elution Volume from 100mAU to 100mAU for Device Articles is shown in FIG. 9.

Table O

[00167] The invention of this application has been described above both generically and with regard to specific embodiments. It will be apparent to those skilled in the art that various modifications and variations can be made in the embodiments without departing from the scope of the disclosure. Thus, it is intended that the embodiments cover the modifications and variations of this invention provided they come within the scope of the appended claims and their equivalents.

Claims

WHAT IS CLAIMED IS:

1 . A composite membrane for an affinity chromatography device comprising: a dry fibri Hated polymer membrane having therein inorganic particles, at least one of the inorganic particles and dry fibril lated polymer membrane having covalently bonded thereto an affinity ligand that reversibly binds to a targeted molecule, wherein the dry fibri Hated polymer membrane has a moisture content of less than or equal to 60% by mass of the dry fibril lated membrane.

2. The composite membrane of claim 1 , wherein the inorganic particles are selected from spherical particles, non-spherical particles, and combinations thereof.

3. The composite membrane of claim 1 or claim 2, wherein the inorganic particles have a spherical shape and a nominal particle size from about 5 microns to about 20 microns.

4. The composite membrane of any one of claims 1 to 3, wherein the inorganic particles have a particle size distribution has a D90/D10 less than or equal to 3.

5. The composite membrane of any one of claims 1 to 4, wherein the dry fibrillated polymer membrane comprises an expanded polytetrafluoroethylene membrane, an expanded modified polytetrafluoroethylene membrane, an expanded tetrafluoroethylene copolymer membrane, or an expanded polyethylene membrane.

6. The composite membrane of any one of claims 1 to 5, wherein the affinity ligand is selected from Protein A, Protein G, Protein L, human Fc receptor protein, antibodies, polysaccharides, oligosaccharides, oligonucleotides and combinations thereof.

7. The composite membrane of any one of claims 1 to 6, wherein the targeted molecule comprises a protein, antibody, viral vector, nucleic acid or combinations thereof.

8. The composite membrane of any one of claims 1 to 7, wherein the dry fibrillated polymer membrane has a wound configuration.

9. The composite membrane of any one of claims 1 to 8 wherein the dry fibrillated polymer membrane has a stacked configuration.

10. The composite membrane of any one of claims 1 to 9, wherein the dry fibrillated polymer membrane is treated for sterilization with ethylene oxide, gamma irradiation, or x-ray irradiation.

11. An affinity chromatography device comprising: a fluid inlet; a fluid outlet fluidly connected to the fluid inlet; a dry fibrillated polymer membrane positioned between the fluid inlet and fluid outlet and containing therein at least one inorganic particle; wherein at least one of: the dry fibrillated polymer membrane has a moisture content less than or equal to 60% by mass of the dry fibrillated polymer membrane prior to drying, or the affinity chromatography device has a moisture content less than or equal to 60% by mass of the affinity chromatography device; and wherein at least one of the dry fibrillated polymer membrane and the inorganic particles has covalently bonded thereto an affinity ligand that reversibly binds to a targeted molecule.

12. The device of claim 11 , comprising a housing encompassing the fluid inlet, the fluid outlet and the dry fibrillated polymer membrane.

13. The device of claim 11 or claim 12, wherein two or more of the dry fibrillated membrane is in the form of a dry stacked membrane assembly positioned between the fluid inlet and fluid outlet.

14. The device of claim 11 or claim 12, wherein the dry fibrillated membrane has a wound membrane configuration comprising the dry fibrillated polymer wound around a core.

15. The device of any one of claims 11 to 14, wherein the targeted molecule comprises a protein, antibody, viral vector, nucleic acid or combinations thereof.

16. The device of any one of claims 11 to 15, wherein the inorganic particles are selected from spherical particles, non-spherical particles, and combinations thereof.

17. The device of any one of claims 11 to 16, wherein the dry fibrillated polymer membrane comprises an expanded polytetrafluoroethylene membrane, an expanded modified polytetrafluoroethylene membrane, an expanded tetrafluoroethylene copolymer membrane, or an expanded polyethylene membrane.

18. The device of any one of claims 11 to 17, wherein the affinity ligand is selected from Protein A, Protein G, Protein L, human Fc receptor protein, antibodies, polysaccharides, oligosaccharides, and combinations thereof.

19. The device of any one of claims 11 to 18, wherein at least one of the dry fibrillated polymer membrane and the affinity chromatography device is treated with ethylene oxide, gamma irradiation or x-ray irradiation.

20. The device of any one of claims 11 to 19, wherein the affinity chromatography device is configured to achieve a dynamic binding capacity (DBC) of at least 35 mg/ml at a residence time of 20 seconds.

21 . The device of any one of claims 11 to 20, wherein the affinity chromatography device is configured to achieve a hydraulic permeability from about 100 (X 10^-12 cm²) to about 2000 (X 1 Q-¹² cm²).

22. The device of any one of claims 11 to 21 , wherein the affinity chromatography device is configured to achieve an elution volume from 100 mAU to 100 mAU of 1 column volume (CV) to 6 column volumes (CV).

23. The device of any one of claims 11 to 22, wherein the affinity chromatography device is configured to achieve a cycling durability of at least 100 cycles at an operating pressure less than 0.3 MPa.

24. Use of the article of any one of claims 11 to 23 to separate the targeted molecule from a fluid stream.

25. A manifold comprising at least two of the affinity chromatography devices of any one of claims 11 to 23 arranged in a parallel configuration.

26. The manifold of claim 25, wherein the manifold is enclosed within a housing.

27. A device comprising a first manifold and a second manifold in a parallel configuration, wherein each of the first manifold and the second manifold includes at least two of the affinity chromatography devices of any one of claims 11 to 23.

28. The device of claim 27, wherein the first manifold and the second manifold are enclosed within a housing.

29. A method of isolating a targeted molecule comprising: providing a chromatography device including: an inlet; an outlet; and a dry fibrillated polymer membrane with a moisture content less than or equal to 60% by mass of the dry fibrillated membrane, the dry fibrillated polymer membrane being positioned between the inlet and the outlet and containing therein at least one inorganic particle; wetting the dry fibri Hated polymer membrane to form a wet polymer membrane; adding an aqueous mixture containing therein a target molecule to the inlet; passing the aqueous mixture through the wet fibrillated polymer membrane where the target molecule is bound to an affinity ligand, and binding the target molecule to an affinity ligand, wherein at least one of the dry fibrillated polymer membrane and the inorganic particle has thereon the affinity ligand.

30. The method of claim 29, comprising treating the dry fibrillated polymer membrane with ethylene oxide to sterilize the dry fibrillated polymer membrane.

31 . The method of claim 30, wherein the treating is conducted in accordance with the Method for Ethylene Oxide (EO) Treatment of Membranes set forth in the Test Method section.

32 The method of claim 29, comprising treating the chromatography device with ethylene oxide, gamma irradiation, or x-ray irradiation.

33 The method of claim 29, comprising treating the chromatography device with ethylene oxide, gamma irradiation, or x-ray irradiation to sterilize the chromatography device.

34. The method of claim 29, comprising treating a non-dry fibrillated membrane to covalently bond an affinity ligand to one or both of the non-dry fibrillated polymer membrane and inorganic particle.

35. The method of claim 29, comprising characterizing the moisture content of the dry fibrillated polymer membrane.

36 The method of claim 35, wherein the characterizing is conducted according to the Method for Determining Moisture Content of Membranes set forth in the Test Methods section.

37 The method of claim 29, comprising characterizing the moisture content of the chromatography device.

38. The method of claim 29, comprising drying a non-dry fibrillated membrane through lyophilization to form the dry fibrillated polymer membrane.

39 The method of claim 38, wherein the drying is conducted according to the Method of Lyophilization of Membranes set forth in the Test Methods section.

40. The method of claim 29, comprising determining the dynamic binding capacity at 10% breakthrough.

41 . The method of claim 40, wherein the determining is conducted according to the Method for Determining the Dynamic Binding Capacity at 10% Breakthrough for Device Articles set forth in the Test Methods section.

42. The method of claim 29, comprising initially wetting the dry chromatography device.

43. The method of claim 42, wherein the initially wetting is conducted according to the Method for Initial Wetting of Dry Device Articles set forth in the Test Methods section.

44. The method of claim 29, comprising determining an elution volume for the chromatography device.

45. The method of claim 44, wherein the determining is conducted according to the Method for Determining Elution Volume from 100mAU to lOOmAU for Device Articles set forth in the Test Methods section.

46. The method of claim 29, comprising determining the dynamic binding capacity at 10% breakthrough for the chromatograph device.

47. The method of claim 46, wherein the determining is conducted according to the Method for Determining the Dynamic Binding Capacity at 10% Breakthrough for Device Articles set forth in the Test Methods section.

48. The method of claim 29, wherein the chromatography device is an affinity chromatography device.

49. The method of claim 29, wherein the affinity ligand is selected from Protein A, Protein G, Protein L, human Fc receptor protein, antibodies, polysaccharides, oligosaccharides, and combinations thereof.

50. The method of claim 29, wherein the inorganic particle are selected from spherical particles, non-spherical particles, and combinations thereof.

51 . The method of claim 29, wherein the dry f ibril lated polymer comprises an expanded polytetrafluoroethylene membrane, an expanded modified polytetrafluoroethylene membrane, an expanded tetrafluoroethylene copolymer membrane, or an expanded polyethylene membrane.

52. An affinity chromatography device comprising: a housing member; an inlet to permit fluid flow into the housing member; an outlet to permit fluid flow out of the housing member and fluidly connected to the inlet; a stacked membrane assembly positioned within the housing member between the fluid inlet and fluid outlet, the stacked membrane assembly comprising: two or more dry fi bril lated polymer membranes in a stacked configuration, each said dry fibrillated polymer membrane containing therein inorganic particles; wherein the dry fibrillated polymer membrane has a moisture content of less than or equal to 60% by mass of the dry fibrillated polymer membrane; and wherein at least one of the dry fibrillated polymer membrane and the inorganic particles has covalently bonded thereto an affinity ligand that reversibly binds to a targeted molecule.

53. The device of claim 52, wherein the targeted molecule comprises a protein, antibody, viral vector, nucleic acid or combinations thereof.

54. The device of claim 51 or claim 52, wherein the inorganic particles are selected from spherical particles, non-spherical particles, and combinations thereof.

55. The device of any one of claims 52 to 54, wherein the inorganic particles have a spherical shape and a nominal particle size from about 5 microns to about 20 microns.

56. The device of any one of claims 52 to 55, wherein a particle size distribution has a D90/D10 less than or equal to 3.

57. The device of any one of claims 52 to 56, wherein the dry fibrillated polymer membrane comprises an expanded polytetrafluoroethylene membrane, an expanded modified polytetrafluoroethylene membrane, an expanded tetrafluoroethylene copolymer membrane, or an expanded polyethylene membrane.

58. The device of any one of claims 52 to 57, wherein the affinity ligand is selected from Protein A, Protein G, Protein L, human Fc receptor protein, antibodies, polysaccharides, oligosaccharides, oligonucleotides and combinations thereof.

59. The device of any one of claims 52 to 58, wherein the device is treated with ethylene oxide, gamma irradiation, or x-ray irradiation.

60. The device of any one of claims 52 to 59, wherein the dry fibril lated polymer membrane has a stacked configuration with two sides and a plurality of edges of the stacked membrane assembly, and wherein the device further comprises a material sealing the edges of the stacked membrane assembly.