Cell filtering devices

Abstract

Devices for filtering liquids are provided herein. More particularly, the devices may be devices for separating biological cells from microcarriers the cells have been grown with, as well as other debris and materials, that are found in the cell culturing liquid. The devices may also be foldable or collapsible.

Description

Attorney Docket No. SP24-301CELL FILTERING DEVICESCROSS REFERENCE TO RELATED APPLICATION

[0001] This application claims the benefit of priority under 35 U.S.C. §119 of U.S. Provisional Application Serial No. 63 / 723,751 filed on November 22, 2024, the content of which is relied upon and incorporated herein by reference in its entirety.FIELD

[0002] The present disclosure relates to devices for filtering liquids, and more particularly to foldable or collapsible devices to separate biological cells from microcarriers and other debris and materials in cell culturing media.BACKGROUND

[0003] Currently existing methods for separating cells grown on microcarriers (tiny substrates that adherent cells bind to and then replicate on) are cumbersome and inefficient because the separation and purification of the cells from the microcarriers requires multiple vessels and fluid exchanges. This problem makes it impractical and expensive to perform large scale cultures with microcarriers, which means that the potentially useful therapies derived from these cultures are either extremely expensive to make or are not practical make despite their usefulness in treating patients. There is therefore a need for improved cell harvesting systems so that harvesting cells that are grown on microcarriers is both more efficient and applicable to large scale cell productions with microcarriers.SUMMARY

[0004] According to some aspects of the present disclosure, microcarrier removal devices are provided. These devices efficiently separate cells from microcarriers, microcarrier debris, extracellular vesicles and unwanted three-dimensional aggregates, and include separating cells from microcarriers that are heterogeneous in size and dimension.

[0005] The present disclosure further relates to foldable or collapsible microcarrier removal devices employing environmentally friendly materials, thus offering sustainability. In certain aspects, the foldable or collapsible devices are particularly well-suited for large-scale microcarrier removal. In one embodiment, the device for filtering liquid comprises a foldable liquid holding chamber / funnel having a lid attached to a base containing a filter media (e.g.,Attorney Docket No. SP24-301 microporous membrane). Such a device may be secured to the top of a secondary container. In another embodiment, the collapsible fdtering device may be used with or without vacuum assistance, as the user could apply positive pressure by pressing or squeezing the flexible top chamber / funnel .

[0006] In another embodiment, the collapsible devices described herein are easy to use (e.g., vacuum pressure may not be required depending on the application), the inclusion of a pleated membrane increases fdtering surface area, the compact nature of the devices frees up valuable laboratory storage space, and they do not require that users alter steps in their routine workflow and best practices in cell culture. In other aspects, the compact design of the collapsible devices of the present disclosure leads to a smaller footprint for shipping to users (i.e., more product per box). Such design results in less void space in waste disposal containers, less material is required construction, less product damage with shipping and handling (removes brittleness factor of top container), and less packaging materials are required.

[0007] In one embodiment, the present disclosure provides a cell separation device configured for separating cells from microcarriers in a liquid, the cell separation device comprising a housing vessel, an inlet port, and an outlet port; and an interior filter unit comprising a physical structure, a first filter membrane and a second filter membrane wherein the first filter membrane is inside the physical structure whereas the second filter membrane is outside the physical structure thereby creating a void space between the first and second filter membranes as defined by the physical structure; wherein the interior filter unit divides an internal space of the housing vessel into three compartments, comprising a first compartment defined by the housing vessel and the first filter membrane, a second compartment defined by a space between the first filter membrane and the second filter membrane, and a third compartment defined by the housing vessel and the second filter membrane.

[0008] In some embodiments the device is single use or multiuse, and recyclable. The first filter membrane may have larger sized pores than those of the second filter membrane. For instance, the first filter membrane comprises pores of about 70 microns to about 150 microns, and the second filter membrane has pores of about 10 microns to about 100 microns. In certain aspects, when the device is configured such that when a microcarrier containing cell solution or suspension enters via the inlet port, the solution or suspension passes the first filter membrane to remove large sized microcarriers, then passes the second filter membrane toAttorney Docket No. SP24-301 remove smaller microcarriers or debris, followed by the twice fdtered solution or suspension reaching the outlet port.

[0009] In further embodiments, the housing vessel comprises a bio- or planet-based material. For example, the bio-based or plant-based material is selected from the group consisting of a thermoplastic starch, polylactic acid (PLA), polyhydroxy fatty acids (PHA), polyethylene terephthalate (PET), monoethylene glycol (MEG), polybutylene succinate (PBS), a polyurethane, a polyolefin, and mixtures of any of the foregoing. In one embodiment, the resin mixture comprises between 20% and 60% bio-based resin and between 40% and 80% petroleum-based resin.

[0010] In other embodiments, the device is foldable to a size smaller than an unfolded size of the device, e.g., the smaller size is at least 25% less than the unfolded size, at least 50% less than the unfolded size, at least 75% less than the unfolded size, or at least 90% less than the unfolded size.

[0011] The present disclosure further provides for a method of separating cells from microcarriers in a liquid, comprising the steps of providing a foldable, cell separation device, unfolding the device from a contracted sate to an expanded state, feeding the microcarrier containing liquid into the inlet port. The foldable, cell separation device comprises (1) a housing vessel, an inlet port, and an outlet port, (2) an interior filter unit comprising a physical structure, a first filter membrane and a second filter membrane; wherein the first filter membrane is inside the physical structure whereas the second filter membrane is outside of the physical structure thereby created avoid space between the first and second filter membranes as defined by the physical structure, (3) one or more folds or pleats to make the device foldable to a size smaller than an unfolded size of the device. The interior filter unit divides an internal space of the housing vessel into three components, comprising a first compartment defined by the housing vessel and the first filter membrane, a second compartment defined by a space between the first filter membrane and the second filter membrane, and a third compartment defined by the housing vessel and the second filter membrane. Once a microcarrier containing liquid is fed into the inlet port, the liquid passes the first filter membrane to remove large sized microcarriers, then passes the second filter membrane to remove smaller microcarriers or debris, followed by the twice filtered liquid reaching the outlet port.Attorney Docket No. SP24-301

[0012] In some embodiments, the contracted state is about 25% less than the expanded state, or about 50% less than the expanded state, or about 75% less than the expanded state, or about 90% less than the expanded state.

[0013] The devices of the present disclosure, in certain embodiments, remove more than 80%, or 85%, or 90%, or 95%, or 99% of the microcarriers, microcarrier debris, and unwanted cell aggregates from the cell culture liquid.

[0014] Additional features and advantages will be set forth in the detailed description which follows, and in part will be readily apparent to those skilled in the art from that description or recognized by practicing the embodiments as described herein, including the detailed description which follows, the claims, as well as the appended drawings.

[0015] It is to be understood that both the foregoing general description and the following detailed description are merely exemplary and are intended to provide an overview or framework to understanding the nature and character of the claims. The accompanying drawings are included to provide a further understanding and are incorporated in and constitute a part of this specification. The drawings illustrate one or more embodiment(s), and together with the description serve to explain principles and operation of the various embodiments.BRIEF DESCRIPTION OF THE DRAWINGS

[0016] The following is a description of the figures in the accompanying drawings, given purely by way of non-limiting example. The figures are not necessarily to scale, and certain features and certain views of the figures may be shown exaggerated in scale or in schematic in the interest of clarity and conciseness.

[0017] FIG. l is a schematic view of a microcarrier removal device, according to some aspects of the present disclosure.

[0018] FIG. 2 is a top view of an interior filter unit 115 illustrated in FIG. 1, according to some aspects of the present disclosure.

[0019] FIG. 3 is a schematic view illustrating the physical structure 120 illustrated in FIG. 1, according to some aspects of the present disclosure.Attorney Docket No. SP24-301

[0020] FIG. 4 is schematic view of a three compartment microcarrier removal device, according to some aspects of the present disclosure.

[0021] FIG. 5 is a schematic view of a foldable three compartment microcarrier removal device, according to some aspects of the present disclosure.

[0022] FIGS. 6A-6D are a series of schematic views of the mechanism of folding of the top container of the filter unit, according to some aspects of the present disclosure.DETAILED DESCRIPTION

[0023] The various aspects and embodiments will now be fully described herein. These aspects and embodiments may, however, be embodied in many different forms and should not be construed as limiting; rather, these embodiments are provided so the disclosure will be thorough and complete, and will fully convey the scope of the present subject matter to those skilled in the art. All publications, patents and patent applications cited herein, whether supra or infra, are hereby incorporated by reference in their entirety.

[0024] Modifications of the disclosure will occur to those skilled in the art and to those who make or use the disclosure. Therefore, it is understood that the embodiments shown in the drawings and described above are merely for illustrative purposes and not intended to limit the scope of the disclosure, which is defined by the following claims, as interpreted according to the principles of patent law, including the doctrine of equivalents.A. Definitions

[0025] Unless defined otherwise, all terms and phrases used herein include the meanings that the terms and phrases have attained in the art, unless the contrary is clearly indicated or clearly apparent from the context in which the term or phrase is used. Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present disclosure, particular methods and materials are now described.

[0026] As used herein the terms “the,” “a,” or “an,” mean “at least one,” and should not be limited to “only one” unless explicitly indicated to the contrary. Thus, for example, referenceAttorney Docket No. SP24-301 to “a component” includes embodiments having two or more such components unless the context clearly indicates otherwise.

[0027] Unless otherwise stated, the use of individual numerical values are stated as approximations as though the values were preceded by the word “about” or “approximately.” Similarly, the numerical values in the various ranges specified in this application, unless expressly indicated otherwise, are stated as approximations as though the minimum and maximum values within the stated ranges were both preceded by the word “about” or “approximately.” In this manner, variations above and below the stated ranges can be used to achieve substantially the same results as values within the ranges. As used herein, the terms “about” and “approximately” when referring to a numerical value shall have their plain and ordinary meanings to a person of ordinary skill in the art to which the disclosed subject matter is most closely related or the art relevant to the range or element at issue. The amount of broadening from the strict numerical boundary depends upon many factors. For example, some of the factors which may be considered include the criticality of the element and / or the effect a given amount of variation will have on the performance of the claimed subject matter, as well as other considerations known to those of skill in the art. As used herein, the use of differing amounts of significant digits for different numerical values is not meant to limit how the use of the words “about” or “approximately” will serve to broaden a particular numerical value or range. Thus, as a general matter, “about” or “approximately” broaden the numerical value. Also, the disclosure of ranges is intended as a continuous range including every value between the minimum and maximum values plus the broadening of the range afforded by the use of the term “about” or “approximately.” Consequently, recitation of ranges of values herein are merely intended to serve as a shorthand method of referring individually to each separate value falling within the range, and each separate value is incorporated into the specification as if it were individually recited herein.

[0028] As used herein, the term “and / or,” when used in a list of two or more items, means that any one of the listed items can be employed by itself, or any combination of two or more of the listed items can be employed. For example, if a composition is described as containing components A, B, and / or C, the composition can contain A alone; B alone; C alone; A and B in combination; A and C in combination; B and C in combination; or A, B, and C in combination.

[0029] As used herein, the terms “foldable” and “collapsible” are used interchangeably.Attorney Docket No. SP24-301

[0030] As used herein, “have,” “having,” “include,” “including,” “comprise,” “comprising” or the like are used in their open ended sense, and generally mean “including, but not limited to.”

[0031] As used herein, “micrometer”, “micron”, and “pm” are used interchangeably.

[0032] “Optional” or “optionally” means that the subsequently described element, component or circumstance may or may not occur, so that the description includes instances where the element, component, or circumstance occurs and instances where it does not.

[0033] In this document, relational terms, such as first and second, top and bottom, and the like, are used solely to distinguish one entity or action from another entity or action, without necessarily requiring or implying any actual such relationship or order between such entities or actions.

[0034] All scientific and technical terms used herein have meanings commonly used in the art unless otherwise specified. The definitions provided herein are to facilitate understanding of certain terms used frequently herein and are not meant to limit the scope of the present disclosure.B. Introduction

[0035] Sterile laboratory media filter devices are used to remove particulates, such as microcarriers, and other contaminates from media, reagents, and buffers used in cell culture compositions. These devices are used in place of alternative methods such as autoclaving, which can lead to degradation of sensitive biological components commonly found in cell culture reagents. Media filter devices are also less capital intensive than autoclaving and can process large volumes of liquid in a reasonable amount of time with limited effort.

[0036] Microcarriers are typically small beads or disks that support cell growth and expansion in bioreactors. They are usually 90-350 microns (pm) in size and are suspended in a medium. Microcarriers are made from a variety of materials, including synthetic materials like dextran, plastic, and glass, and natural materials like cellulose, gelatin, and collagen. The past decades have witnessed increasing use of microcarriers for scale-up of adherent cell culture, in particular, stem cells comprising human mesenchymal stem cells (hMSCs). This has been driven by the increased use of high doses of hMSCs (e.g., from 105to 109cells per patient)Attorney Docket No. SP24-301 in autologous or allogenic cell therapy, as well as the ability of microcarriers to be adopted with stirred tank culture systems. Because hMSCs are developed to be implanted, transplanted, or infused into human patients, efficient purification processes are essential for safe and effective delivery.

[0037] Microcarrier removal is part of the cell purification process. Because most microcarriers used today are not biodegradable, they must be removed before the cells are implanted, transplanted, or infused into patients. A variety of devices and techniques have been used to separate cells from microcarriers during the cell clarification process. These include sedimentation using conical or inclined settlers, centrifugation, acoustic resonance, spin filtration, microfiltration, and inertial-based filtration. However, many of these devices and techniques are not single use and require sophisticated equipment, routine maintenance and between-use cleaning and sterilization.

[0038] Single use devices also have disadvantages as plastic, single-use media filter devices are just one example of the calculable plastic waste that is generated in scientific laboratories annually. One paper estimated that 5.5 million metric tons of plastic are generated from scientific laboratories each year. Uriba MA et al., Labs should cut plastic waste too, Nature 528: 479 (2015) (DOI: 10.1038 / 528479c). Current disposable filter units are comprised of a hard plastic top chamber / funnel that adds to plastic waste and that take up valuable space during shipping, storage, and disposal. They also typically require vacuum assistance to apply negative pressure in the receiving container to encourage filtration.

[0039] Others have attempted to reduce the amount of plastic used in filter products. For example, EMD Millipore introduced Stericup® E and SteriTop® E funnel-less filter units claim to reduce plastic and packaging. However, these products still require secondary containers for the filtering process, which places the burden on the user to supply the primary and secondary containers for filter processing. Such an approach does not lead to less plastic waster or packaging reduction on a net basis.

[0040] Other collapsible laboratory products have been introduced as space savings solutions, but without environmental sustainability benefits. For instance, MTC Bio introduced the Bio Bucket™, which is a collapsible laboratory ice bucket or portable liquid container. The collapsible bucket provides space saving solutions and comes at a significantly lower cost (about $50 / unit) compared to traditional laboratory buckets (about $100 / unit). Yet, not onlyAttorney Docket No. SP24-301 does the Bio Bucket™ lack environmental advantages, but its lifetime may be significantly shorter due to its construction and material composition. Foldable bottles made of bio-based materials (e.g., DiFOLD Origami) are available, but lack means for filtering cell culture and have not been adapted or designed for laboratory settings and uses.

[0041] There is a need, therefore, for filtering devices to be composed of less plastic, derived from more Earth-friendly components, require less transportation costs and carbon emissions, and are easy to use by customers.

[0042] The microcarrier removal devices disclosed herein are designed to be efficient in separating cells from microcarriers, microcarrier debris, and unwanted cell aggregates, including in separating cells from microcarriers that are heterogenous in size and dimension. The devices may be of rigid structure (e.g., plastic or non-plastic compositions), or foldable / collapsible. Microcarrier removal devices that are foldable / collapsible may be packaged and / or stored in a much smaller amount of space compared to non-flexible, non- collapsible devices.

[0043] Additional features and advantages will be set forth in the detailed description which follows and will be apparent to those skilled in the art from the description or recognized by practicing the embodiments as described in the following description, together with the claims and appended drawings.C. Filter Devices

[0044] The microcarrier removal devices described herein employ one or more filters to separate cells from microcarriers. The filtering devices may be constructed in a variety of ways, for example, as described in U.S. Pat. No. 11,788,051 and U.S. Pat. Pub. No. 2024 / 0002778, both commonly assigned to Coming, Inc. and incorporated herein by reference. For instance, in one embodiment, the microcarrier filtering devices use a filter membrane that is a vertical or inclined porous mesh or membrane (or other filter configuration) to capture microcarriers within a vessel, where they can easily be washed and treated to remove the cells from the microcarriers. As used herein, a “filter membrane” refers to the filter for the microcarriers, which may be a mesh, a membrane, or other configuration that has pores sized to allow desired materials such as liquid and harvested cells to flow through the pores but prevents microcarriers from flowing through. The mesh (or membrane) may be pleated or folded to increase the filtering surface area. The incline or vertical positioning of the filterAttorney Docket No. SP24-301 membrane within the device prevents the microcarriers or unwanted cell aggregates from blocking the pores of the mesh to enhance fluid flow and cell separation. The design of the device enables the liquid (e.g., cell culture medium, buffered saline) to flow over the inclined or vertical fdter membrane such that the microcarriers or cell aggregates roll off the fdter membrane and can still be retained in the device. In other embodiments, the microcarrier fdtering devices use a horizontal filter membrane that is a porous mesh or membrane (or other filter configuration) to capture microcarriers within a vessel, where they can also easily be washed and treated to remove the cells from the microcarriers.

[0045] The device can be made as an open or closed system. The device may be filled by pouring, gravity, perfusion, vacuum, or by pressurization of the source vessel. In certain embodiments, no vacuum is necessary for any of the steps in the process to remove the microcarrier materials or to filter the cells. The device can be structured so that it is easily integrated into a perfusion bioreactor system. The device is self-contained and after isolating the microcarriers or cell aggregates on one side of the filter membrane, inverting the device permits further manipulation. The cells may be washed and released from the microcarriers in the device, and then collected by perfusing them out of the device. The device is self-contained and can potentially be used to transport cells for therapy to the site of administration.

[0046] The device can be many shapes and scaled in size as needed. The device may be a rigid vessel or a flexible vessel. The device may have an outer housing for stabilization or it can be made without an outer housing as desired. The device can be manufactured using all the conventional methods for forming and assembling devices from plastic materials or other types of materials. To enhance desired attributes the device may be coated or treated. The filter membrane can be made from a polymer, metal, glass, or composites thereof, and the pore sizes can range, for example, from about 12-200 microns.

[0047] The housing vessels or containers and the filter membranes of the present disclosure may be made from a variety of plastic and / or non-plastic materials. In one aspect, they are made from resin comprising a thermoplastic copolyester (TPC) resin. The resin may be 100% bio- or plant-based, for example, from rapeseed or other plant material. Alternatively, the TPC resin may be a mixture of bio- or plant-based and petroleum-based, for example, between 20% to 60% bio-based resin and between 40% and 80% petroleum-based resin. One example of a petroleum-based component of a mixed TPC resin is Amitel resin manufactured by Envalior (Netherlands). Another suitable material includes petroleum-based polyethyleneAttorney Docket No. SP24-301 terephthalate (PET), a polyester that has a semi-crystalline form when stable. It is recyclable and shows resistance to impact, moisture, alcohols, and solvents. Other suitable materials include petroleum-based polyurethane (PU) and petroleum-based polyolefins such as petroleum-based polyethylene (PE) and petroleum-based polypropylene (PP), among others known to those of ordinary skill in the art.

[0048] Bioplastics suitable for use in the filtering devices include: (a) thermoplastic starch, which can be made from com, wheat, potato, and tapioca; (b) polylactic acid (PLA), which is a bio-based aliphatic polyester that is obtained by fermenting sugar and starch; (c) polyhydroxy fatty acids (PHF), which are thermoplastic polyesters produced by bacteria or fungi from starch and used for medical applications such as sutures, slings, bone plates, and skin substitutes; (d) polyethylene furanoate (PEF), a high-performance plastic similar to PET; (e) bio-based PET, such as Coca-Cola’s PlantBottle™ material, which is a fully recyclable bio- PET made of bio-based monoethylene glycol (MEG); (f) polybutylene succinate (PBS); (g) bio-based polyurethanes (PU); or (h) bio-based polyolefins, such as bio-based polyethylene (PE) and bio-based polypropylene (PP).

[0049] The seals, ports, and other components of the filtering devices may be comprised of silicone or other inert, non-reactive materials. Preferably the devices are 100% BPA (bisphenol A) / BPS (bisphenol-S)-free, and do not contain lead or toxins. In other aspects, the devices are recyclable, including closed-loop recyclable, which means they can be recycled without losing their properties. Further, the foldable or collapsible devices are durable and can withstand multiple folds without damaging the folding creases or other means for folding or collapsing.

[0050] Embodiments describing the vertical or inclined filter membranes will now be described. Referring to FIGS. 1-3, in one embodiment, a microcarrier removal device 100 is provided. Removal device 100 comprises a housing vessel 110, an interior filter unit 115, an inlet port 160, and an outlet port 150. The interior filter unit 115 further comprises a physical structure 120, a first filter membrane 130, a second filter membrane 140, wherein the first filter membrane 130 is placed inside the physical structure 120, while the second filter membrane 140 is place outside the physical structure 120. In this configuration, there is a void space between the first and second filter membrane, as defined by the physical structure 120.Attorney Docket No. SP24-301

[0051] The interior filter unit 115 divides the internal space of the housing vessel 110 into three components, wherein the first compartment is defined by the center of the housing vessel 110 and the first filter membrane 130, the second compartment is defined by the space between the first filter membrane 130 and the second filter membrane 140, and the third compartment is defined by the interior wall of the housing vessel 110 and the second filter membrane 140. In this configuration, when a microcarrier cell mixture solution enters the device 100 via the inlet port 160, the solution will pass the first filter membrane 130 to remove large sized microcarriers, then the second filter membrane 140 to remove smaller microcarriers or microcarrier debris, finally reaching the outlet port 150.

[0052] The first filter membrane 130 has larger sized pores than those of the second filter membrane 140. For instance, the first filter membrane 130 has pores of about 70 micrometers to about 150 micrometers, where the second filter membrane 140 has pores of about 30 micrometers to about 100 micrometers. The filter membrane can be made of suitable plastics, such as nylon, polypropylene, and polyester, among others known to those of ordinary skill in the art.

[0053] Referring to FIGS. 2-3, the physical structure 120 further comprises a top ring structure 116, and an open frame structure 117. The edge of the top ring structure 116 is in tight seal with the inner wall of the housing vessel 110. The sealing can be achieved via adhesive, direct thermal fusion, or ultrasonic welding driven fusion, among other types of sealing known to those of ordinary skill in the art. The first filter membrane 130 can directly at least partially fuse onto the inner wall surface of the open frame structure 117, while the second filter membrane 140 can directly at least partially fuse onto the outer wall surface of the open frame structure 117.

[0054] An embodiment with membrane filters in a horizonal configuration will now be described. Referring to FIG. 4, there is a three compartment microcarrier removal device 200. The microcarrier removal device 200 comprises a housing vessel 210 that is separated into three compartments by a first filter membrane 240, and a second filter membrane 250. When a cell microcarrier mixture solution enters the device 200 via the inlet port 200, the solution will enter the first compartment, pass the first filter membrane 240, enter the second compartment, pass the second filter membrane 250. The instance, the first filter membrane 240 has pore sizes of about 70 micrometers to about 150 micrometers, whereas the second filter membrane 250 has pore sizes of about 30 micrometers to about 100 micrometers. The first filter membraneAttorney Docket No. SP24-301 can be made of suitable plastics, such as nylon, polypropylene, and polyester, among others known to those of ordinary skill in the art.D. Collapsible / Foldable Filter Devices

[0055] The filter devices of the present disclosure may be collapsable or foldable. More specifically, the housing and / or any supports of the filter device may be collapsable or foldable. Such filter devices may have one or more filter membranes.

[0056] FIGS. 6A-6D illustrate, in one embodiment, the folding mechanism of the top container of the filter unit. The container comprises a flexible polyester with the appropriate strategic folds to allow for folding yet retain the structural integrity when open and holding volumes of liquid. In certain embodiments, a tab, strap, or other locking means could be added to the bottom of the container, that can be positioned or secured around the top of the container when it is in a collapsed state, to ensure the security of the folded container during shipping. The locking means (including a tab or strap) can be any known to those of ordinary skill in the art, including a removable or non-removable elastic strap that goes around the entirety of the folded container, a tab with one or more snap closures that secures the bottom of the folded container to the top of the folded container, or a strap or tab with hook and loop closures that secures the top and bottom portions of the container when the container is in its collapsed state.

[0057] The collapsible / foldable containers comprise folds on the sidewalls of the collapsible / foldable portion of the container (e.g., the housing 110 of the filter unit shown in FIGS. 6A-6D). In some embodiments, the inflection points of the folds may be approximately horizontal to the bottom portion of the container. FIG. 5 illustrates a container having inflection points of the fold that are approximately horizontal to the bottom portion of the container. In one specific embodiment, a container comprises sidewalls and the sidewalls of the container comprise at least one fold with inflection points that are approximately horizontal to the bottom portion of the container. In one further embodiment, the container may further comprise two filters, each in an approximately horizontal configuration, and the sidewalls of the container comprise at least three folds with inflect points that are approximately horizontal to the bottom portion on the container.

[0058] In other embodiments, the inflection points of the fold may be angular (not approximately horizontal) to the bottom portion of the container. FIG. 6 illustrates a container having inflection points of the fold that are angular to the bottom portion of the container. InAttorney Docket No. SP24-301 one specific embodiment, a container comprises sidewalls and the sidewalls of the container comprise at least two folds with inflection points that are angular to the bottom portion of the container. In one further embodiment, the container may further comprise at least one filter.

[0059] In certain aspects, the devices comprise one or more foldable sections of different shapes and sizes. They can either be single use or reusable. The single use products can easily be collapsed and disposed of more effectively than non-collapsible products and so significantly reduce the waste volume. The structural advantages of the foldable devices include:• They are self-supporting and stable in unfolded positions, e.g., on laboratory benches• They can have smooth and round surfaces• They are compact in folded positions• They can comprise both cylindrical and / or conical surfaces• They can operate also in a partly folded state, giving variability in shape and size

[0060] In one specific embodiment, referring to FIG. 5, a foldable three compartment microcarrier removal device 300 is disclosed. The microcarrier removal device 300 comprises a housing vessel 310 that is separated into three compartments by a first horizontal filter membrane 340, and a second horizontal filter membrane 350. When in an operating position, a cell microcarrier mixture solution enters the device 300 via the inlet port 320, the solution will enter the first compartment, pass the first horizontal filter membrane 340, enter the second compartment, pass the second horizontal filter membrane 350, and enter the third compartment, finally exiting the device via outlet port 330. The first horizontal filter membrane 340 has pores sized larger than those of the second horizontal filter membrane 350. For instance, the first horizontal filter membrane has pore sizes of between 70 micrometers and 150 micrometers, whereas the second horizontal filter membrane 350 has pore sizes of between 30 micrometers and 100 micrometers. The first horizontal filter membrane 240 can be made of suitable plastics, such as nylon, polypropylene, and polyester, among others known to those of ordinary skill in the art. The housing vessel 310 is foldable such that when being packaged, the whole device can be stored under compressed form and only occupy such smaller dimensions.E. Illustrative AspectsAttorney Docket No. SP24-301

[0061] Aspect 1. A cell separation device is provided that is configured for separating cells from microcarriers. The cell separation device comprises (1) a housing vessel, an inlet port, and an outlet port; and (2) an interior filter unit comprising a physical structure, a first filter membrane and a second filter membrane. The first filter membrane is inside the physical structure whereas the second filter membrane is outside the physical structure thereby creating a void space between the first and second filter membranes as defined by the physical structure. The interior filter unit divides an internal space of the housing vessel into three compartments, comprising a first compartment defined by the housing vessel and the first filter membrane, a second compartment defined by a space between the first filter membrane and the second filter membrane, and a third compartment defined by the housing vessel and the second filter device.

[0062] Aspect 2. The cell separation device of aspect 1, wherein the device is a single use device or a multiuse device.

[0063] Aspect 3. The cell separation device of any one of aspects 1-2, wherein the device is recyclable.

[0064] Aspect 4. The cell separation device of any one of aspects 1-3, wherein the filter membranes are selected from the group consisting of nylon, polypropylene, polystyrene, polyethylene, polyethylene terephthalate, polylactic acid, polyhydroxyalkanoate, and combinations thereof.

[0065] Aspect 5. The cell separation device of any one of aspects 1-4, wherein the first filter membrane has larger sized pores than those of the second filter membrane.

[0066] Aspect 6. The cell separation device of aspect 5, wherein the first filter membrane comprises pores of about 70 microns to about 150 microns, and the second filter membrane has pores of about 10 microns to about 100 microns.

[0067] Aspect 7. The cell separation device of any one of aspects 1-6, wherein the device is foldable to a size smaller than an unfolded size of the device.

[0068] Aspect 8. The cell separation device of aspect 7, wherein the smaller is size is between 15% and 35% less than the unfolded size.

[0069] Aspect 9. The cell separation device of aspect 7, wherein the smaller size is between 40% and 60% less than the unfolded size.Attorney Docket No. SP24-301

[0070] Aspect 10. The cell separation device of aspect 7, wherein the smaller size is between 65% and 85% less than the unfolded size.

[0071] Aspect 11. The cell separation device of aspect 7, wherein the smaller size is greater than or equal to 85% less than the unfolded size.

[0072] Aspect 12. The cell separation device of any one of aspects 1-11, wherein the device is configured such that when a microcarrier containing cell solution or suspension enters via the inlet port, the solution or suspension passes the first filter membrane to remove large sized microcarriers, then passes the second filter membrane to remove smaller microcarriers or debris, and then reaches the outlet.

[0073] Aspect 13. A foldable, cell separation device is provided that is configured for separating cells from microcarriers. The cell separation device comprises (1) a housing vessel, an inlet port, and an outlet port; and (2) an interior filter unit comprising a physical structure, a first filter membrane and a second filter membrane. The first filter membrane is inside the physical structure whereas the second filter membrane is outside the physical structure thereby creating a void space between the first and second filter membranes as defined by the physical structure. The interior filter unit divides an internal space of the housing vessel into three compartments, comprising a first compartment defined by the housing vessel and the first filter membrane, a second compartment defined by a space between the first filter membrane and the second filter membrane, and a third compartment defined by the housing vessel and the second filter membrane. The device is foldable to a size smaller than an unfolded size of the device.

[0074] Aspect 14. The cell separation device of aspect 13, wherein the housing vessel comprises a bio-based material.

[0075] Aspect 15. The cell separation device of aspect 14, wherein the bio-based material is selected from the group consisting of a thermoplastic startch, polylactic acid (PLA), polyhydroxy fatty acids (PHF), bio-based polyethylene terephthalate (PET), monoethylene glycol (MEG), or polybutylene succinate (PBS), and combinations of the foregoing.

[0076] Aspect 16. The cell separation device of aspect 13, wherein the housing vessel comprises a thermoplastic copolyester (TPC) resin mixture of bio-based resin and petroleumbased resin.Attorney Docket No. SP24-301

[0077] Aspect 17. The cell separation device of aspect 16, wherein the resin mixture comprises between 20% and 60% bio-based resin and between 40% and 80% petroleum-based resin.

[0078] Aspect 18. The cell separation device of any one of aspects 13-17, wherein the small size is between 15%-35% less than the unfolded size.

[0079] Aspect 19. The cell separation device of any one of aspects 13-17, wherein the smaller size is between 40% and 60% less than the unfolded size.

[0080] Aspect 20. The cell separation device of any one of aspects 13-17, wherein the small size is between 65% and 85% less than the unfolded size.

[0081] Aspect 21. The cell separation device of any one of aspects 13-17, wherein the smaller size is greater than or equal to 85% less than the unfolded size.

[0082] Aspect 22. A foldable, cell separation device is provided that is configured for separating cells from microcarriers. The cell separation device comprises (1) a housing vessel, an inlet port, and an outlet port; (2) an interior filter unit comprising a physical structure, a first filter membrane and a second filter membrane; and (3) a top ring structure and an open frame structure. The first filter membrane is inside the physical structure whereas the second filter membrane is outside the physical structure thereby creating a void space between the first and second filter membranes as defined by the physical structure. An edge of the top ring structure in full or partial sealable engagement with an inner wall of the housing vessel. The second filter membrane fully or partially fuses onto an outer wall surface of the open frame structure The interior filter unit divides an internal space of the housing vessel into three compartments, comprising a first compartment defined by the housing vessel and the first filter membrane, a second compartment defined by a space between the first filter membrane and the second filter membrane, and a third compartment defined by the housing vessel and the second filter membrane. The device is foldable to a size smaller than an unfolded size.

[0083] Aspect 23. The cell separation device of aspect 22, wherein the housing vessel comprises bio-based material.

[0084] Aspect 24. The cell separation device of aspect 23, wherein the bio-based material is selected from the group consisting of a thermoplastic startch, polylactic acid (PLA),Attorney Docket No. SP24-301 polyhydroxy faty acids (PHF), bio-based polyethylene terephthalate (PET), monoethylene glycol (MEG), or polybutylene succinate (PBS), and combinations of the foregoing.

[0085] Aspect 25. The cell separation device of aspect 22, wherein the housing vessel comprises a thermoplastic copolyester (TPC) resin mixture of bio-based resin and petroleumbased resin.

[0086] Aspect 26. The cell separation device of aspect 25, wherein the resin mixture comprises between 20% and 60% bio-based resin and between 40% and 80% petroleum-based resin.

[0087] Aspect 27. The cell separation device any one of aspects 22-26, wherein the small size is between 15%-35% less than the unfolded size.

[0088] Aspect 28. The cell separation device of any one of aspects 22-26, wherein the smaller size is between 40% and 60% less than the unfolded size.

[0089] Aspect 29. The cell separation device of any one of aspects 22-26, wherein the small size is between 65% and 85% less than the unfolded size.

[0090] Aspect 30. The cell separation device of any one of aspects 22-26, wherein the smaller size is greater than or equal to 85% less than the unfolded size.

[0091] Aspect 31. A method of separating cells from microcarriers in a liquid is provided, comprising the steps of: (a) providing a foldable, cell separation device; (b) unfolding the device from a contracted state to an expanded state; and (c) feeding the microcarrier containing liquid into the inlet port, the liquid passing through the first filter membrane to remove large size microcarriers, then passing through the second filter membrane to remove smaller microcarriers or debris, the reaching the outlet port. The cell-separation device comprises (1) a housing vessel, an inlet port, and an outlet port; (2) an interior filter unit comprising a physical structure, a first filter and a second membrane filter; and (3) one or more folds or pleats to make the device foldable to a size smaller than an unfolded size of the device. The first filter membrane is inside the physical structure whereas the second membrane filter is outside the physical structure thereby creating a void space between the first and second filter membranes as defined by the physical structure. The interior filter unit divides an internal space of the housing vessel into three components, comprising a first compartment defined by theAttorney Docket No. SP24-301 housing vessel and the first filter membrane, a second compartment defined by a space between the first filter membrane and the second filter membrane, and a third compartment defined by the housing vessel and the second filter membrane.

[0092] Aspect 32. The cell separation device of aspect 31, wherein the housing vessel comprises a bio-based material.

[0093] Aspect 33. The cell separation device of aspect 32, wherein the bio-based material is selected from the group consisting of a thermoplastic startch, polylactic acid (PLA), polyhydroxy fatty acids (PHF), bio-based polyethylene terephthalate (PET), monoethylene glycol (MEG), or polybutylene succinate (PBS), and combinations of the foregoing.

[0094] Aspect 34. The cell separation device of aspect 33, wherein the resin mixture comprises between 20% and 60% bio-based resin and between 40% and 80% petroleum-based resin.

[0095] Aspect 35. The cell separation device of any one of aspects 31-34, wherein the small size is between 15%-35% less than the unfolded size.

[0096] Aspect 36. The cell separation device of any one of aspects 31-34, wherein the smaller size is between 40% and 60% less than the unfolded size.

[0097] Aspect 37. The cell separation device of any one of aspects 31-34, wherein the small size is between 65% and 85% less than the unfolded size.

[0098] Aspect 38. The cell separation device of any one of 31-34, wherein the smaller size is greater than or equal to 85% less than the unfolded size.

[0099] While the present disclosure includes a limited number of embodiments, those skilled in the art, having benefit of this disclosure, will appreciate that other embodiments can be devised which do not depart from the scope of the present disclosure.

Claims

Attorney Docket No. SP24-301CLAIMSWhat Is Claimed Is:

1. A cell separation device configured for separating cells from microcarriers, the cell separation device comprising: a housing vessel, an inlet port, and an outlet port; and an interior filter unit comprising a physical structure, a first filter membrane and a second filter membrane, wherein the first filter membrane is inside the physical structure whereas the second filter membrane is outside the physical structure thereby creating a void space between the first and second filter membranes as defined by the physical structure; wherein the interior filter unit divides an internal space of the housing vessel into three compartments, comprising a first compartment defined by the housing vessel and the first filter membrane, a second compartment defined by a space between the first filter membrane and the second filter membrane, and a third compartment defined by the housing vessel and the second filter device.

2. The device of claim 1, wherein the device is single use or multiuse.

3. The device of claim 1, wherein the device is recyclable.

4. The device of claim 1, wherein the filter membranes are selected from the group consisting of nylon, polypropylene, polystyrene, polyethylene, polyethylene terephthalate, polylactic acid, polyhydroxyalkanoate, and combinations thereof.

5. The device of claim 1, wherein the first filter membrane has larger sized pores than those of the second filter membrane.

6. The device of claim 1, wherein the first filter membrane comprises pores of about 70 microns to about 150 microns, and the second filter membrane has pores of about 10 microns to about 100 microns.

7. The device of claim 1, wherein the device is foldable to a size smaller than an unfolded size of the device.

8. The device of claim 7, wherein the smaller is size is between 15% and 35% less than the unfolded size.Attorney Docket No. SP24-3019. The device of claim 7, wherein the smaller size is between 40% and 60% less than the unfolded size.

10. The device of claim 7, wherein the smaller size is between 65% and 85% less than the unfolded size.

11. The device of claim 7, wherein the smaller size is greater than or equal to 85% less than the unfolded size.

12. The device of claim 1, wherein the device is configured such that when a microcarrier containing cell solution or suspension enters via the inlet port, the solution or suspension passes the first filter membrane to remove large sized microcarriers, then passes the second filter membrane to remove smaller microcarriers or debris, and then reaches the outlet.

13. A foldable, cell separation device configured for separating cells from microcarriers, the cell separation device comprising: a housing vessel, an inlet port, and an outlet port; and an interior filter unit comprising a physical structure, a first filter membrane and a second filter membrane wherein the first filter membrane is inside the physical structure whereas the second filter membrane is outside the physical structure thereby creating a void space between the first and second filter membranes as defined by the physical structure; wherein the interior filter unit divides an internal space of the housing vessel into three compartments, comprising a first compartment defined by the housing vessel and the first filter membrane, a second compartment defined by a space between the first filter membrane and the second filter membrane, and a third compartment defined by the housing vessel and the second filter membrane; and wherein the device is foldable to a size smaller than an unfolded size of the device.

14. The device of claim 13, wherein the housing vessel comprises a bio-based material.

15. The device of claim 14, wherein the bio-based material is selected from the group consisting of a thermoplastic startch, polylactic acid (PLA), polyhydroxy fatty acids (PHF), bio-based polyethylene terephthalate (PET), monoethylene glycol (MEG), or polybutylene succinate (PBS), and combinations of the foregoing.Attorney Docket No. SP24-30116. The device of claim 13, wherein the housing vessel comprises a thermoplastic copolyester (TPC) resin mixture of bio-based resin and petroleum -based resin.

17. The device of claim 16, wherein the resin mixture comprises between 20% and 60% bio-based resin and between 40% and 80% petroleum-based resin.

18. The device of claim 13, wherein the small size is between 15%-35% less than the unfolded size.

19. The device of claim 13, wherein the smaller size is between 40% and 60% less than the unfolded size.

20. The device of claim 13, wherein the small size is between 65% and 85% less than the unfolded size.

21. The device of claim 13, wherein the smaller size is greater than or equal to 85% less than the unfolded size.

22. A foldable, cell separation device configured for separating cells from microcarriers, the cell separation device comprising: a housing vessel, an inlet port, and an outlet port; an interior filter unit comprising a physical structure, a first filter membrane and a second filter membrane wherein the first filter membrane is inside the physical structure whereas the second filter membrane is outside the physical structure thereby creating a void space between the first and second filter membranes as defined by the physical structure; and a top ring structure and an open frame structure, wherein an edge of the top ring structure in full or partial sealable engagement with an inner wall of the housing vessel and wherein the second filter membrane fully or partially fuses onto an outer wall surface of the open frame structure; wherein the interior filter unit divides an internal space of the housing vessel into three compartments, comprising a first compartment defined by the housing vessel and the first filter membrane, a second compartment defined by a space between the first filter membrane and the second filter membrane, and a third compartment defined by the housing vessel and the second filter membrane; andAttorney Docket No. SP24-301 wherein the device is foldable to a size smaller than an unfolded size.

23. The device of claim 22, wherein the housing vessel comprises bio-based material.

24. The device of claim 23, wherein the bio-based material is selected from the group consisting of a thermoplastic startch, polylactic acid (PLA), polyhydroxy fatty acids (PHF), bio-based polyethylene terephthalate (PET), monoethylene glycol (MEG), or polybutylene succinate (PBS), and combinations of the foregoing.

25. The device of claim 22, wherein the housing vessel comprises a thermoplastic copolyester (TPC) resin mixture of bio-based resin and petroleum -based resin.

26. The device of claim 25, wherein the resin mixture comprises between 20% and 60% bio-based resin and between 40% and 80% petroleum-based resin.

27. The device of claim 22, wherein the small size is between 15%-35% less than the unfolded size.

28. The device of claim 22, wherein the smaller size is between 40% and 60% less than the unfolded size.

29. The device of claim 22, wherein the small size is between 65% and 85% less than the unfolded size.

30. The device of claim 22, wherein the smaller size is greater than or equal to 85% less than the unfolded size.

31. A method of separating cells from microcarriers in a liquid, comprising the steps of:(a) providing a foldable, cell separation device, the cell-separation device comprising: a housing vessel, an inlet port, and an outlet port; an interior fdter unit comprising a physical structure, a first filter and a second membrane filter, wherein the first filter membrane is inside the physical structure whereas the second membrane filter is outside the physical structure thereby creating a void space between the first and second filter membranes as defined by the physical structure; wherein the interior filter unit divides an internal space of the housing vessel into three components, comprising a first compartment defined by the housing vessel and theAttorney Docket No. SP24-301 first filter membrane, a second compartment defined by a space between the first filter membrane and the second filter membrane, and a third compartment defined by the housing vessel and the second filter membrane; and one or more folds or pleats to make the device foldable to a size smaller than an unfolded size of the device;(b) unfolding the device from a contracted state to an expanded state;(c) feeding the microcarrier containing liquid into the inlet port, the liquid passing through the first filter membrane to remove large size microcarriers, then passing through the second filter membrane to remove smaller microcarriers or debris, the reaching the outlet port.

32. The device of claim 31, wherein the housing vessel comprises a bio-based material.

33. The device of claim 32, wherein the bio-based material is selected from the group consisting of a thermoplastic startch, polylactic acid (PLA), polyhydroxy fatty acids (PHF), bio-based polyethylene terephthalate (PET), monoethylene glycol (MEG), or polybutylene succinate (PBS), and combinations of the foregoing.

34. The device of claim 33, wherein the resin mixture comprises between 20% and 60% bio-based resin and between 40% and 80% petroleum-based resin.

35. The device of claim 31, wherein the small size is between 15%-35% less than the unfolded size.

36. The device of claim 31, wherein the smaller size is between 40% and 60% less than the unfolded size.

37. The device of claim 31, wherein the small size is between 65% and 85% less than the unfolded size.

38. The device of claim 31, wherein the smaller size is greater than or equal to 85% less than the unfolded size.

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