Microcavity cell culture vessel with diverter comprising angled surface

Abstract

A cell culture vessel includes a vessel body including a first end wall, a second end wall opposite the first end wall, and a pair of side walls extending between the first end wall and the second end wall. The first end wall, the second end wall, and the pair of side walls define a perimeter of the cell culture chamber. An inner fill wall extends inwardly into the cell culture chamber from the first end wall and the pair of side walls. The inner filler wall has an angled inner surface having an outer edge at the first end wall that is angularly offset from an inner terminal edge of the angled inner surface. The diverter extends between the pair of sidewalls along the inclined inner surface at least partially transverse to the filling direction. The diverter provides a passage between the diverter and the pair of sidewalls through which liquid flows along the diverter during a filling operation.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of priority under 35 U.S.C. §119 to U.S. Provisional Application Serial No. 63 / 411,210, filed on September 29, 2022, the contents of which are hereby relied upon and incorporated herein by reference in their entirety. Technical Field

[0002] The present specification relates generally to cell culture containers for growing cells and, more particularly, to microcavity cell culture containers having flow diverters that restrict and control the flow of liquid down the inclined surface of the cell culture container. background

[0003] In general, three-dimensional (3D) cell culture can be better suited to simulating the environment of natural tissues and organs than two-dimensional (2D) cell culture grown in monolayer.Compared to 2D cells grown in monolayer, cells grown in 3D cell culture can adhere to other deposited cells in a three-dimensional environment and form spheroids or cell aggregates, thereby establishing more natural interactions between cells.This natural arrangement of cells provides a flexible configuration similar to natural tissue.It is expected that an accurate example of tissue microenvironment is provided.In order to improve accuracy in experimental studies for developing disease therapy, it is expected that the therapy is developed in 3D culture rather than 2D culture, because 3D culture is closer to the environment where the developed drug will eventually be administered.

[0004] However, when transporting the cell culture container or during the culture medium replacement, the formed spheroids or cell aggregates are vulnerable to damage. Transporting 3D cell culture containers generally causes the liquid contained therein to move unexpectedly, resulting in turbulence in the container. In the 3D cell culture process, turbulence may also occur when adding liquid culture medium to the container or removing liquid culture medium from the container. Turbulence may cause spheroids or cell aggregates to shake or shift in their respective microcavities in the 3D culture container, because the cells in the 3D cell culture are not attached to any surface of the container. If the spheroid is displaced from its respective microcavity, the formed spheroid may be attached to other formed spheroids, resulting in the loss of uniformity and size of the spheroid in the container. Therefore, it is necessary to stabilize the movement of liquid in the 3D spheroid culture container. Summary of the invention

[0005] According to one embodiment, a cell culture container includes a container body, the container body including a first end wall, a second end wall opposite to the first end wall, and a pair of side walls extending between the first end wall and the second end wall. The first end wall, the second end wall and the pair of side walls define the perimeter of a cell culture chamber. An inner filling wall extends inwardly from the first end wall and the pair of side walls into the cell culture chamber. The inner filling wall has an inclined inner surface, the inclined inner surface having an outer edge at the first end wall, the outer edge being offset at an angle from the inner terminal edge of the inclined inner surface. A diverter extends between the pair of side walls along the inclined inner surface at least partially transverse to a filling direction. The diverter provides a passage between the diverter and the pair of side walls, through which liquid flows along the diverter during a filling operation.

[0006] According to another embodiment of the present disclosure, a method for forming a cell culture container is provided. The method includes forming a container body with a single integral component, so that the container body includes a first end wall, a second end wall opposite to the first end wall, and a pair of side walls extending between the first end wall and the second end wall. The first end wall, the second end wall and the pair of side walls define the perimeter of the cell culture chamber. The inner filling wall extends inwardly from the first end wall and the pair of side walls into the cell culture chamber. The inner filling wall has an inclined inner surface, which has an outer edge at the first end wall, and the outer edge is offset at an angle from the inner terminal edge of the inclined inner surface. The diverter extends between the pair of side walls along the inclined inner surface at least partially transverse to the filling direction. The diverter provides a passage between the diverter and the pair of side walls, and during the filling operation, the liquid flows along the diverter through the passage. The protective carrier is formed separately from the container body. The protective carrier includes a plate, which is configured and sized to cover the cell culture surface of the container body.

[0007] Other features and advantages of the cell culture container described herein are presented in the following detailed description, some of which are readily apparent to those skilled in the art from the description or recognized by practicing the embodiments described herein, including the following detailed description, claims, and drawings.

[0008] It should be understood that both the foregoing general description and the following detailed description describe various embodiments and are intended to provide an overall review or framework for understanding the nature and characteristics of the claimed subject matter. The included drawings provide a further understanding of the various embodiments, and the drawings are incorporated into and constitute a part of this specification. The drawings illustrate the various embodiments described herein and, together with the description, are used to explain the principles and operation of the claimed subject matter. BRIEF DESCRIPTION OF THE DRAWINGS

[0009] Figure 1According to one or more embodiments shown and described herein, a cell culture container is shown that includes a container body having a flow diverter and a protective carrier;

[0010] Figure 2 According to one or more embodiments shown and described herein, a Figure 1 A side cross-sectional view of a container body;

[0011] Figure 3 According to one or more embodiments shown and described herein, the container body is shown along Figure 2 An end cross-sectional view of line 3-3;

[0012] Figure 4 According to one or more embodiments shown and described herein, a Figure 1 A top perspective view of a protective carrier;

[0013] Figure 5 According to one or more embodiments shown and described herein, Figure 4 A bottom perspective view of a protective carrier;

[0014] Figure 6 According to one or more embodiments shown and described herein, a Figure 1 a side perspective view of a portion of a cell culture substrate of a cell culture vessel of the invention; and

[0015] Figure 7 According to one or more embodiments shown and described herein, a Figure 1 Cell culture vessels. DETAILED DESCRIPTION

[0016] Reference will be made in detail below to various embodiments of a cell culture container in which a flow diverter is positioned, examples of which are shown in the accompanying drawings. Whenever possible, the same reference numerals will be used throughout the drawings to refer to the same or similar parts. Directional terms used herein - e.g., up, down, right, left, front, back, top, bottom, far, and near - are only with reference to the drawings in which they are drawn and are not intended to imply an absolute orientation.

[0017] Herein, ranges may be expressed as starting from "about" one particular value and / or ending at "about" another particular value. When such a range is expressed, another embodiment includes starting from the one particular value and / or ending at the other particular value. Similarly, when a value is expressed as an approximation using the antecedent "about," it is understood that the particular value constitutes another embodiment. It is also understood that the endpoints of each range are significant both in relation to the other endpoint and independently of the other endpoint.

[0018] Unless otherwise expressly stated, any method described herein should not be construed as requiring that its steps be performed in a specific order, or requiring that any apparatus have a particular orientation. Thus, if a method claim does not actually recite the order in which its steps are to be followed, or any apparatus does not actually recite the order or orientation of its components, or if the claims or specification do not otherwise specifically state that the steps are limited to a specific order, or if a specific order or orientation of apparatus components is not recited, then no order or orientation should be inferred in any respect. This applies to any possible non-expressive basis for interpretation, including: logical issues involving the arrangement of steps, operational flow, order of components, or orientation of components; issues of obvious meaning derived from grammatical organization or punctuation; and issues of the number or type of embodiments described in the specification.

[0019] As used herein, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise. Thus, for example, reference to "an" component includes aspects having two or more such components unless the context clearly dictates otherwise.

[0020] Liquid in a cell culture container with an open, wide volume can move freely and uncontrollably throughout the open area. If the cell culture container is moved or transported, the movement can cause turbulence in the liquid, causing the liquid to "slosh" and possibly spill. Unlike cells cultured in two-dimensional (2D) culture that are attached to the culture surface of the container, cells (e.g., spheroids) in 3D culture may not be attached to the culture surface and therefore may be easily sloshed and may be displaced from the microcavity in which they are being cultured. This turbulence and displacement of spheroids from the microcavity may cause the spheroids to be lost. Since displaced spheroids may eventually settle in the microcavity with other spheroids, the size of the spheroids may become uneven, resulting in an undesirable loss of uniformity of the spheroids.

[0021] Further, a certain degree of skill is required to fill the microcavities with culture medium without trapping air, and to replace the culture medium after spheroid formation. Trapped air can cause cells to be unable to settle in the microcavities. Moreover, since the cells in the cell culture container grow in 3D, the container can accommodate many more cells than the cells that are usually accommodated in the 2D cell growth surface area. The increased number of cells may require more frequent replacement of culture medium or accommodating larger fluid volumes to meet the metabolic needs of the cells.

[0022] Embodiments described herein relate to a cell culture container, which includes a filling structure that can be used to control the filling of the cell culture container with a liquid culture medium. The filling structure can also control the movement of the culture medium after the cell culture container is filled. The cell culture container includes a container body, the container body including a first end wall, a second end wall opposite to the first end wall, and a pair of side walls extending between the first end wall and the second end wall. When the bottom and the cover are connected to the container body, the first end wall, the second end wall and the pair of side walls form the perimeter of the cell culture chamber. The container body includes a neck extending outward from the first end wall in a filling direction toward the opening. The neck includes an inclined filling surface, the inclined filling surface having an outer edge at the opening, the outer edge and the inner edge of the inclined filling surface are offset at an angle at the first end wall. The inner filling wall extends inward from the first end wall and the pair of side walls into the cell culture chamber. The inner filling wall has an inclined inner surface, the inclined inner surface having an outer edge at the first end wall, the outer edge and the inner terminal edge of the inclined inner surface are offset at an angle. The flow divider extends between the pair of side walls along the inclined inner surface at least partially transverse to the filling direction. The flow divider has opposite ends that terminate inside the pair of side walls to form a passage through which liquid flows along the flow divider during a filling operation.

[0023] refer to Figure 1 and 2 , cell culture container 10( Figure 1 ) is an embodiment comprising a container body 12 ( Figure 2 ) of the micro-chamber flask. The container body 12 includes a first end wall 14 and a second end wall 16 opposite the first end wall 14. Side walls 18 and 20 extend between the first end wall 14 and the second end wall 16. In some embodiments, the first end wall 14 and the second end wall 16 or portions thereof are substantially parallel, and a pair of side walls 18 and 20 or portions thereof are substantially parallel, thereby forming a slightly rectangular perimeter 22 of the cell culture chamber 24. Although a rectangular perimeter 22 is illustrated, any suitable perimeter shape may be used.

[0024] The container body 12 includes a neck 26 that extends outwardly from the first end wall 14 in a filling direction toward an opening 28. The opening 28 provides an entry point for liquid culture medium to enter the cell culture container 10 and flow into the cell culture chamber 24 during a filling operation. The opening 28 also provides an exit point for liquid culture medium to leave the cell culture chamber 24 during an emptying operation. The neck 26 can optionally be configured to mate with the lid 27, for example, using threads, a snap fit, or any suitable releasable connection.

[0025] The neck 26 includes an inclined filling surface 30 that forms part of the neck 26, the inclined filling surface 30 having an outer edge 32 at the opening 28 that is angularly offset from an inner edge 34 of the inclined filling surface 30 at the first end wall 14. An inner filling wall 36 extends inwardly from the first end wall 14 and the pair of side walls 18 and 20 into the cell culture chamber 24. The inner filling wall 36 has an inclined filling surface 38 that has an outer edge 40 at the first end wall 14 that is angularly offset from an inner terminal edge 42 of the inclined inner surface 38. In some embodiments, the inclined inner surface 38 is an extension of the inclined filling surface 30 of the neck 26, wherein the inclined inner surface 38 and the inclined filling surface 30 intersect and are disposed when the cell culture container 10 is in a position such as Figure 1 and 2 In the case of the horizontal incubation position shown, it extends at the same angle θ to the horizontal (e.g., greater than or equal to 10 degrees, greater than or equal to 15 degrees, greater than or equal to 20 degrees, between 10 degrees and 20 degrees, between 15 degrees and 20 degrees).

[0026] The cell culture container 10 also includes a lid 44 that covers a viewing surface 46 of the container body 12. The lid 44 includes a rim 47 that engages the edges 48 of the first and second end walls 14, 16 and the side walls 18, 20 to provide an interlocking connection. The lid 44 can be formed of a clear material, such as a clear or translucent plastic, so that the contents within the cell culture chamber 24 can be visually inspected.

[0027] The cell culture container 10 can include a cell culture substrate 50 positioned along an opposing cell culture surface 52 of the container body 12. The cell culture substrate 50 can include a surface 54 that includes a plurality of microcavities 56 that are sized and shaped to receive at least one cell or spheroid therein. Thus, the cell culture surface 54 is a cell culture area configured to promote growth and development of cells in the cell culture chamber 24.

[0028] refer to Figure 2 The container body 12 forms a frame 58 at the cell culture surface 52 of the container body 12, and its size is configured to receive the cell culture substrate 50 (see Figure 1). Specifically, the frame 58 can be formed at one edge 61 thereof, the edge 61 being located between a ledge 62 extending outwardly from the inner fill wall 36 and an inner terminal edge 42 of the inner fill wall 36, the ledge 62 extending outwardly from the inner fill wall 36 away from the cell culture chamber 24, the inner terminal edge 42 being offset below the inclined inner surface 38. An opposite edge 63 of the frame 58 can be formed between another ledge 64 and an edge 66 of the second end wall 16, the other ledge 64 extending outwardly from the second end wall 16 away from the cell culture chamber 24. The edges of the frame 58 can also be formed in a similar manner along the side walls 18 and 20 (including ledges 67 and 69) (see Figure 3 ). During the incubation process, the cell culture substrate 50 can be held within the frame 58. As can be seen, the edge 68 of the cell culture substrate 50 can be offset below the inner end edge 42 of the inner filling wall 36.

[0029] Specific reference Figure 3 , the flow divider 70 extends along the inclined inner surface 38. The flow divider 70 extends between the pair of side walls 18 and 22 at least partially transverse to the filling direction. The flow divider 70 is formed as a partial height wall because the height H1 of the flow divider is less than the height H2 of the cell culture chamber 24 (e.g., less than or equal to 50%, less than or equal to 60%, less than or equal to 70%, less than or equal to 80%, less than or equal to 90%). The flow divider 70 has opposite ends 72 and 74 that terminate inside the pair of side walls 18 and 20 to form passages 76 and 78 through which liquid culture medium flows along the flow divider 70 during filling and emptying operations.

[0030] The diverter 70 extends in the width direction along the inclined inner surface 38. In some embodiments, the diverter 70 is integrally formed with the inner filling wall 36 as a single integral piece of the inner filling wall 36. In other embodiments, the diverter 70 can be formed separately and connected to the inclined inner surface 38. The diverter 70 can contact and connect with the inclined inner surface 38 along its entire length to guide the liquid culture medium through the passages 76 and 78. Although the diverter 70 is illustrated as a rectangular cross-sectional shape, it can be formed in other shapes, such as a curved shape, a triangle, etc. Moreover, the diverter 70 can extend in other directions through the width of the inclined inner surface 28. For example, the diverter 70 can extend at an oblique angle to the filling direction.

[0031] The flow divider 70 may extend only partially along the width W of the angled inner surface 38 between the ends 72 and 74 to provide passages 76 and 78. For example, the flow divider 70 may extend no more than 99% of the width W, for example, no more than 95%, for example, no more than 90%, for example, no more than 85%, for example, no more than 80%, for example, no more than 75%, for example, no more than 70%, for example, no more than 65%, for example, no more than 60%, for example, no more than 55%, for example, no more than 50%, for example, no more than 45%, for example, no more than 40%, for example, no more than 35%, for example, no more than 30% of the width W. In some embodiments, the flow divider 70 extends at least about 50% and no more than about 90% of the width, for example, about 80% of the width W.

[0032] refer to Figure 4 , the protective carrier 80 is a rigid plate that can protect the gas permeable microcavities of the cell culture substrate 50 during transportation and can also serve as a protective carrier for the cell culture container 10 during use for cell culture. The protective carrier 80 can include a rigid plate 82 that is slightly larger than the occupied area of ​​the cell culture container 10. An outer edge 84 can extend around the perimeter of the plate 82 and can be configured and sized to extend around the protrusions 62, 64, 67, 69 of the inner fill wall 36 and the second end wall 16 and the side walls 18 and 20. Standoff ribs 86 are provided at the corners of the protective carrier 80. The standoff ribs 86 extend along both the plate 82 and the outer edge 84 to provide gaps and air gaps between the protective carrier 80 and the cell culture substrate 50. Reference Figure 5 Along the underside of the plate 82, a stacking groove or rib 88 may be provided. The stacking groove or rib 88 is provided to mate with a corresponding stacking rib or groove 90 ( Figure 1 )Cooperate.

[0033] Reference now Figure 6 , gas permeability is a property that contributes to a 3D cell culture environment. By allowing gas permeability within the microcavity 100 of the cell culture container, the cell culture growth medium may need to be replaced less frequently and cell growth may be promoted. The geometry and formation of the microcavity container are unique because they are formed by a gas permeable substrate 50 having micrometer-sized pores 100 (also referred to as microcavities). Such microcavity containers enjoy gas permeability due to the thickness of the microcavity substrate 50, wherein the gas permeability occurs because the microcavity substrate can be formed from a very thin polystyrene manufacturing material having a thickness of about 28 microns to about 72 microns. Although gas permeability may be an advantage for culturing cell aggregates, the thinness of the microcavity substrate material makes the microcavity cell culture container 10 vulnerable to damage during transportation and use.

[0034] Each microcavity 100 may include an inner cavity having a rounded bottom 102 that is non-adhesive to cells. Thus, the microcavity container as described herein is a cell culture device that facilitates 3D cell culture by allowing cells seeded into the microcavities to self-assemble or attach to each other to form spheroids in each microcavity. The microcavities may be shallow and allow cell culture medium to cover the spheroids, organoids, or 3D cell aggregates in all cavities at once to facilitate manual operation.

[0035] In one embodiment, the top surface 104 of the microcavity 100 may be recessed close to the sidewalls 18 and 20 ( Figure 3 ) at the bottom of the microcavity. A separate microcavity can hold a small amount of culture medium. A separate microcavity can have any suitable size. For example, the diameter or width of a separate microcavity can be in the range of about 500 microns to about 5 millimeters. The depth of a separate microcavity can be in the range of about 500 microns to about 6 millimeters. In some embodiments, the depth of a separate microcavity can be about 500 microns to about 650 microns. In some embodiments, the depth of a separate microcavity can be about 1.6 millimeters. Excess culture medium can be added to the microcavity container so that a spheroid, organoid, or 3D cell aggregate does not need to rely solely on a small amount of culture medium in a separate microcavity.

[0036] In some embodiments, the microcavity substrate 50 can have a wavy cross-sectional shape or a cross-sectional shape that approximates a sine wave. In such an embodiment, the bottom of the microcavity hole 100 is rounded (e.g., hemispherical), the diameter of the sidewall increases from the bottom to the top of the hole, and the boundary or limit between the holes is rounded. Thus, the top of the microcavity hole does not terminate at a right angle. In some embodiments, the width of the hole is greater than the width of the limit between adjacent holes. Such an embodiment allows a greater number of holes in a given area of ​​the culture surface.

[0037] In some embodiments, a plurality of microcavities are arranged in a hexagonal close-packed pattern. In some embodiments, each microcavity includes a rounded bottom. In some embodiments, each microcavity is configured so that cells cultured in a microcavity container form a three-dimensional (3D) cell aggregate. In some embodiments, the inner surface of the microcavity substrate has no adhesion to cells. In some embodiments, the inner surface of the microcavity substrate includes a cell non-adhesive surface coating comprising a perfluoropolymer, an olefin, a lipid, an agarose, a nonionic hydrogel, a polyether, a polyol, a polymer that inhibits cell attachment, or a combination thereof. In some embodiments, the cell non-adhesive surface coating comprises an ultra-low attachment (ULA) surface coating.

[0038] The microcavity substrate, microcavity container, and diverter can be formed of the same material or similar materials. In some embodiments, the microcavity substrate can be molded or formed separately from the rest of the microcavity container and bonded by thermal bonding, ultrasonic welding, or any other plastic connection method. The construction materials of the microcavity container, microcavity substrate, and / or baffle can include "plastic" polymers, copolymers, or polymer blends. Non-limiting examples include silicone rubber, polystyrene, polypropylene, polyethylene, polyethylene terephthalate, polymethylpentene, polycarbonate, polymethyl methacrylate, styrene-ethylene-butadiene-styrene, other such polymers, or combinations thereof. In some embodiments, the microcavity substrate is formed of polydimethylsiloxane (PDMS), polymethylpentene, (poly) 4-methylpentene (PMP), polyethylene (PE), polystyrene (PS), polypropylene, polyethylene terephthalate, polycarbonate, polymethyl methacrylate, styrene-ethylene-butadiene-styrene, silicone rubber or copolymer, ethylene vinyl acetate, polysulfone, polytetrafluoroethylene, poly(styrene-butadiene-styrene), or a combination thereof. Any suitable construction method can be used to form the microcavity substrate, microcavity container, and shunt, such as non-limiting examples including injection molding, thermoforming, 3D printing, or any other method suitable for forming plastic parts.

[0039] In some embodiments, the protective carrier 80 is formed of a polymer, a metal, or a glass. In some embodiments, the polymer comprises polystyrene, polypropylene, polyethylene, polyethylene terephthalate, polymethylpentene, polycarbonate, polymethyl methacrylate, styrene-ethylene-butadiene-styrene, other such polymers, or combinations thereof. In some embodiments, the metal comprises aluminum, stainless steel, zinc, or combinations thereof. In some embodiments, the glass comprises borosilicate glass. In some embodiments, the protective carrier is formed of a recyclable material. In some embodiments, the protective carrier is formed of a biodegradable material. In some embodiments, the protective carrier is opaque. In some embodiments, the protective carrier is translucent.

[0040] refer to Figure 1-3In use, a plurality of cells may be deposited in the cell culture container 10 such that the cell culture chamber 24 may be used to contain cells within a plurality of microcavities 100 of the cell culture surface 110 of the cell culture substrate 50. First, the microcavity surface may need to be pre-wetted. Liquid culture medium may be provided through the neck 26, along the inclined filling surface 30, and then flow through the inclined inner surface 38 of the inner filling wall 36. The liquid culture medium may then impact the diverter 70, which diverts the liquid culture medium along its length toward the channels 76 and 78, so that the liquid culture medium flows more slowly as it approaches the cell culture surface 110. The liquid culture medium may enter the microcavity 100 without assistance. If the liquid does not enter the microcavity, trapped air may appear as opaque areas on the cell culture surface 110. If the microcavity 100 has trapped air, the suspended cells may not settle into these microcavities 100 after the filling process. As the cell culture substrate 50 receives a plurality of cells along the cell culture surface 110, the development of the cells is facilitated by exposing the cell culture surface 110 to various nutrients and growth fluids during the liquid culture medium filling operation.

[0041] The microcavity can be deep enough to allow for gentle culture medium changes, but not too deep to make it difficult to recover the spheroids when needed. During culture medium changes, the cell culture container 10 can be kept in an incubation position to inhibit spheroid loss. The inclined inner surface 38 and the flow divider 70 can promote gentle flow of culture medium from the cell culture container 10.

[0042] During the medium replacement step, lifting the second end wall 16 by three to four degrees can push the liquid medium toward the diverter 70 and the inclined inner surface 38 to achieve complete liquid removal, and can slow down the flow rate of new liquid added to the cells in the culture container 10 to reduce splashing. When aspirating and dispensing liquid, it is advantageous that the lifting angle of the second end wall 16 does not exceed 10 degrees to inhibit liquid from splashing and wetting the cover 27. Figure 7 , the protective carrier 80 or an item of similar height (e.g., between 6 mm and 7 mm) can be used to lift the second end wall 16 during the medium exchange step. Specifically, the protrusions 67 and 69 can include stacking notches 114, which can be used as stop features to capture the outer edge 84 of the protective carrier 80 and place the cell culture container at the appropriate angle. To remove the spent medium, the tip of the pipette can be placed against the diverter 70 to aspirate the medium. To change the medium, the tip of the pipette can be placed against the diverter 70 to slowly add fresh medium to the cell culture container 10. Once the medium exchange is completed, the protective carrier 80 can be removed and the cell culture container 10 can be placed in a flat incubation position.

[0043] The cell culture container includes a shunt located in each cell culture chamber of the microcavity cell culture container, and the system for microcavity cell culture includes a shunt located in each cell culture chamber of the microcavity cell culture container, which reduces the amount of movement of the liquid culture medium in the cell culture chamber. The shunt can also improve filling by controlling the flow of liquid and reducing the trapped air in the microcavity. The diverter can be formed (e.g., molded) with the container body so that the shunt and the container body are formed as a single integral piece of material. The shunt can also provide a position where the tip of the pipette can be placed against it and used to remove the liquid culture medium from the cell culture chamber and distribute the liquid culture medium to the cell culture chamber, which can improve the filling efficiency and the repeatability between the filling and emptying operations.

[0044] Embodiments may be described with reference to the following numbered clauses, wherein preferred features are presented in the dependent clauses:

[0045] Item 1: A cell culture container, comprising: a container body, the container body comprising: a first end wall; a second end wall opposite to the first end wall; a pair of side walls extending between the first end wall and the second end wall, the first end wall, the second end wall and the pair of side walls defining the perimeter of the cell culture chamber; an inner filling wall extending inwardly from the first end wall and the pair of side walls into the cell culture chamber, the inner filling wall having an inclined inner surface, the inclined inner surface having an outer edge at the first end wall, the outer edge being angularly offset from the inner terminal edge of the inclined inner surface; a diverter extending between the pair of side walls along the inclined inner surface at least partially transverse to the filling direction, the diverter providing a passage between the diverter and the pair of side walls, through which liquid flows along the diverter during a filling operation.

[0046] Item 2: The cell culture container of Item 1, wherein the entire length of the flow divider is connected to the inclined inner surface of the inner filling wall.

[0047] Item 3: The cell culture container of Item 2, wherein the flow diverter is formed as an integral, unitary component of the inner fill wall.

[0048] Item 4: The cell culture container of any one of Items 1-3, wherein the flow diverter has opposite ends that terminate inside the pair of side walls to provide a passageway.

[0049] Clause 5: The cell culture container of any one of clauses 1-4, further comprising a cover configured and dimensioned to cover a viewing surface of the container body.

[0050] Clause 6: The cell culture container of any one of clauses 1-5, further comprising a protective carrier comprising a plate configured and dimensioned to cover the cell culture surface of the container body.

[0051] Clause 7: The cell culture container of Clause 6, wherein the protective carrier comprises a rim extending around the perimeter of the plate, the rim being configured and dimensioned to extend along the first end wall, the end side wall, and the pair of side walls.

[0052] Clause 8: The cell culture container of Clause 6 or 7, wherein the protective carrier comprises spacer ribs arranged and configured to be located between the plate and the container body to provide a space between the plate and the container body.

[0053] Item 9: The cell culture container of any of Items 1-8, wherein the diverter extends at least about 50% and no more than about 90% of the width of the sloped interior surface between the ends of the diverter.

[0054] Item 10: A cell culture container according to any one of items 1 to 9, wherein the container body further comprises a neck extending outwardly from the first end wall to the opening in a filling direction, wherein the neck comprises an inclined filling surface having an outer edge at the opening that is angularly offset from an inner edge of the inclined filling surface at the first end wall.

[0055] Item 11: The cell culture container of Item 10, wherein the inclined filling surface and the sloped inner surface intersect.

[0056] Item 12: A method for forming a cell culture container, the method comprising: forming a container body from a single integral component, such that the container body comprises: a first end wall; a second end wall opposite the first end wall; a pair of side walls extending between the first end wall and the second end wall, the first end wall, the second end wall and the pair of side walls defining the perimeter of the cell culture chamber; an inner filling wall extending inwardly from the first end wall and the pair of side walls into the cell culture chamber, the inner filling wall having an inclined inner surface, the inclined inner surface having an outer edge at the first end wall, the outer edge being angularly offset from an inner terminal edge of the inclined inner surface; and a diverter extending between the pair of side walls along the inclined inner surface at least partially transverse to a filling direction, the diverter providing a passage between the diverter and the pair of side walls, through which liquid flows along the diverter during a filling operation; and forming a protective carrier independently of the container body, the protective carrier comprising a plate configured and sized to cover a cell culture surface of the container body.

[0057] Clause 13: The method of clause 12, wherein the entire length of the diverter is connected to the inclined inner surface of the inner fill wall.

[0058] Clause 14: The method of clause 12 or 13, wherein the diverter has opposing ends that terminate inboard of the pair of side walls to provide a passageway.

[0059] Clause 15: The method of any of Clauses 12-14, further comprising forming a cover configured to cover a viewing surface of the container body.

[0060] Clause 16: The method of any of clauses 12-15, wherein the protective carrier comprises a rim extending around a perimeter of the panel, the rim being configured and dimensioned to extend along the first end wall, the end side wall, and the pair of side walls.

[0061] Clause 17: The method of any of clauses 12-16, wherein the protective carrier comprises a spacer rib arranged and configured to be located between the plate and the container body to provide a space between the plate and the container body.

[0062] Clause 18: The method of any of clauses 12-17, wherein the flow splitter extends at least about 50% and no more than about 90% of the width of the beveled inner surface between the ends of the flow splitter.

[0063] Clause 19: The method of any of clauses 12-18, wherein the container body further comprises a neck extending outwardly from the first end wall to the opening in a filling direction, the neck comprising an inclined filling surface having an outer edge at the opening that is angularly offset from an inner edge of the inclined filling surface at the first end wall.

[0064] Clause 20: The method of clause 19, wherein the inclined fill surface and the sloped interior surface intersect.

[0065] It is obvious to those skilled in the art that various modifications and changes can be made to the embodiments described herein without departing from the spirit and scope of the claimed subject matter. Therefore, this specification is intended to cover the modifications and changes of the various embodiments described herein, as long as these modifications and changes are within the scope of the attached claims and their equivalents.

Claims

1. A cell culture container, comprising: A container body, the container body comprising: a first end wall; a second end wall opposite to the first end wall; a pair of side walls extending between the first end wall and the second end wall, the first end wall, the second end wall and the pair of side walls defining a perimeter of the cell culture chamber; an inner fill wall extending inwardly from the first end wall and the pair of side walls into the cell culture chamber, the inner fill wall having a sloped inner surface having an outer edge at the first end wall that is angularly offset from an inner terminal edge of the sloped inner surface; and A flow divider extends at least partially transverse to the filling direction along an inclined inner surface between the pair of side walls, the flow divider providing a passage between the flow divider and the pair of side walls through which liquid flows along the flow divider during a filling operation.

2. The cell culture container according to claim 1, wherein The entire length of the flow divider is connected to the inclined inner surface of the inner filling wall.

3. The cell culture container according to claim 2, wherein: The flow divider is formed as an integral, unitary component of the inner fill wall.

4. The cell culture container according to claim 1, wherein The flow divider has opposite ends that terminate inside the pair of side walls to provide a passageway.

5. The cell culture container of claim 1, further comprising a cover configured and dimensioned to cover a viewing surface of the container body.

6. The cell culture container of claim 1, further comprising a protective carrier comprising a plate configured and dimensioned to cover a cell culture surface of the container body.

7. The cell culture container according to claim 6, wherein: The protective carrier includes a rim extending around the perimeter of the plate, the rim being configured and dimensioned to extend along the first end wall, the end side wall, and the pair of side walls.

8. The cell culture container according to claim 6, wherein: The protective carrier includes a spacer rib arranged and configured to be located between the plate and the container body to provide a space between the plate and the container body.

9. The cell culture container according to claim 1, wherein: The flow divider extends at least about 50% and no more than about 90% of the width of the beveled inner surface between the ends of the flow divider.

10. The cell culture container according to claim 1, wherein: The container body also includes a neck extending outwardly from the first end wall to the opening in a filling direction, the neck including an inclined filling surface having an outer edge at the opening that is angularly offset from an inner edge of the inclined filling surface at the first end wall.

11. The cell culture container according to claim 10, wherein: The sloped fill surface intersects the angled interior surface.

12. A method for forming a cell culture container, the method comprising: The container body is formed as a single integral component such that the container body comprises: a first end wall; a second end wall opposite to the first end wall; a pair of side walls extending between the first end wall and the second end wall, the first end wall, the second end wall and the pair of side walls defining a perimeter of the cell culture chamber; an inner fill wall extending inwardly from the first end wall and the pair of side walls into the cell culture chamber, the inner fill wall having a sloped inner surface having an outer edge at the first end wall that is angularly offset from an inner terminal edge of the sloped inner surface; and a flow divider extending at least partially transverse to the filling direction along an inclined inner surface between the pair of side walls, the flow divider providing a passage between the flow divider and the pair of side walls through which liquid flows along the flow divider during a filling operation; and A protective carrier is formed separately from the container body, the protective carrier comprising a plate configured and dimensioned to cover the cell culture surface of the container body.

13. The method of claim 12, wherein: The entire length of the flow divider is connected to the inclined inner surface of the inner filling wall.

14. The method of claim 12, wherein: The flow divider has opposite ends that terminate inside the pair of side walls to provide a passageway.

15. The method of claim 12, further comprising forming a cover configured to cover a viewing surface of the container body.

16. The method of claim 12, wherein: The protective carrier includes a rim extending around the perimeter of the plate, the rim being configured and dimensioned to extend along the first end wall, the end side wall, and the pair of side walls.

17. The method of claim 12, wherein: The protective carrier includes a spacer rib arranged and configured to be located between the plate and the container body to provide a space between the plate and the container body.

18. The method of claim 12, wherein: The flow divider extends at least about 50% and no more than about 90% of the width of the beveled inner surface between the ends of the flow divider.

19. The method of claim 12, wherein: The container body also includes a neck extending outwardly from the first end wall to the opening in a filling direction, the neck including an inclined filling surface having an outer edge at the opening that is angularly offset from an inner edge of the inclined filling surface at the first end wall.

20. The method of claim 19, wherein: The sloped fill surface intersects the angled interior surface.

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