Patent Publication Number: US-2021189314-A1

Title: Cell culture apparatuses with manifolds including column structures

Description:
CROSS-REFERENCE 
     This application claims the benefit of and priority to U.S. Provisional Application No. 62/582,086, titled “Cell Culture Apparatuses with Manifolds Including Column Structures,” filed Nov. 6, 2017, the details of which are incorporated herein by reference in their entirety. 
    
    
     FIELD 
     The present disclosure relates to apparatuses for culturing cells and, in particular, cell culture apparatuses with manifolds including column structures. 
     BACKGROUND 
     Many types of cell culture articles are constructed to provide stacked or stackable units for culturing cells. For example, T-flasks are typically made to have flat top and bottom surfaces that allow T-flasks to be stacked, providing space savings. Some modified T-flasks have multiple parallel culture surfaces within the flask to reduce time and effort associated with filling and emptying. Other culture apparatuses are multi-component assemblies having a plurality of parallel or stacked culture surfaces. Manipulating stacked cell culture devices can be challenging. These devices must be filled, usually on their sides, and then turned so that the cell culture surfaces are horizontal. It can be challenging to fill and manipulate these large devices, which can be heavy when filled with liquid. In addition, it is important to maintain the integrity and sterility of these devices. 
     An exemplary cell culture article is Corning&#39;s HYPERStack™ system. The HYPERStack™ system includes multiple modules formed of individual stackette layers that can be interconnected by flexible tubes that connect to tube connectors. The modules are interconnected for filling and emptying the HYPERStack™ system. Valves and other devices may be used to control fluid flow into and out of the HYPERStack™ system. The use of these valves and other devices can be cumbersome and provide potential leak locations. There is a need for improved cell culture articles with increased capacity and improved flow control features. In addition, it is desirable to reduce the risk of leaks and contamination, minimize wasteful use of expensive media and valuable cells, in cell culture articles that are easier to maneuver, fill and empty. 
     BRIEF SUMMARY 
     In a first aspect, a cell culture apparatus, comprises: a cell culture module comprising multiple cell culture chambers; and a manifold that connects the multiple cell culture chambers together along a side of the cell culture module, the manifold comprising a side wall base structure connected to the side of the cell culture module and a column structure that is formed as a monolithic part of the side wall base structure, the column structure defining a fluid flow pathway through the manifold and to inlets to the cell culture chambers to allow filling and emptying of the cell culture chambers of liquid medium. 
     According to a second aspect, there is provided the cell culture apparatus of aspect 1, wherein the cell culture apparatus comprises multiple cell culture modules, the manifold comprises manifold segments, each manifold segment is associated with one of the cell culture modules. 
     According to a third aspect, there is provided the cell culture apparatus of aspect 2, wherein each manifold segment comprises a side wall base structure segment and a column structure segment that is formed as a monolithic part of the side wall base structure segment. 
     According to a fourth aspect, there is provided the cell culture apparatus of aspect 3, wherein the column structure segments of adjacent manifold segments are interconnected and in fluid communication. 
     According to a fifth aspect, there is provided the cell culture apparatus of aspect 4, wherein the column structure segments of adjacent manifold segments are interconnected by a sealing ring. 
     According to a sixth aspect, there is provided the cell culture apparatus of aspect 5, wherein the sealing ring seals an interface between the column structure segments of adjacent manifold segments. 
     According to a seventh aspect, there is provided the cell culture apparatus of aspect 5 or 6, wherein the sealing ring comprises relatively thick portions that are separated by a relatively thin portion in the form of a notch that defines an area of increased flexibility compared to the thick portions. 
     According to an eighth aspect, there is provided the cell culture apparatus of any of aspects 5-7, wherein each column structure segment comprises a shroud structure, wherein adjacent shroud structures face each other thereby forming a partial enclosure that extends about an entire periphery of the sealing ring. 
     In a ninth aspect, A method of forming a cell culture apparatus, the method comprising: stacking one cell culture module on another cell culture module, the cell culture modules comprising multiple cell culture chambers; and connecting the cell culture chambers using a manifold, the manifold comprising a side wall base structure and a column structure that is formed as a monolithic part of the side wall base structure, the column structure defining a fluid flow pathway through the manifold and to the cell culture chambers to allow filling and emptying of the cell culture chambers of liquid medium. 
     According to a tenth aspect, there is provided the method of aspect 9, wherein the manifold comprises manifold segments, the method comprising connecting each manifold segment to one of the cell culture modules. 
     In an eleventh aspect, there is provided the method of aspect 10, wherein each manifold segment comprises a side wall base structure segment and a column structure segment that is formed as a monolithic part of the side wall base structure segment. 
     In a twelfth aspect, there is provided the method of aspect 11 comprising connecting the column structure segments of adjacent manifold segments such that the column segments of the adjacent manifold segments are in fluid communication. 
     In a thirteenth aspect, there is provided the method of aspect 12 comprising connecting the column structure segments of adjacent manifold segments by a sealing ring. 
     In a fourteenth aspect, there is provided the method of aspect 13, wherein sealing an interface between the column structure segments of adjacent manifold segments using the sealing ring. 
     In a fifteenth aspect, there is provided the method of aspect 13 or 14, wherein the sealing ring comprises relatively thick portions that are separated by a relatively thin portion in the form of a notch that defines an area of increased flexibility compared to the thick portions. 
     In a sixteenth aspect, there is provided the aspect of any of aspects 13-15, wherein each column structure segment comprises a shroud structure, wherein adjacent shroud structures face each other thereby forming a partial enclosure that extends about an entire periphery of the sealing ring. 
     In a seventeenth aspect, a cell culture apparatus, comprises: at least two cell culture modules, each cell culture module comprising at least two cell culture chambers; and a manifold that connects the at least two cell culture manifolds together, the manifold comprising multiple manifold segments, each manifold segment comprising a side wall base structure segment and a column structure segment that is formed as a monolithic part of the side wall base structure segment, the column structure segments connected together and defining a fluid flow pathway through the manifold and to the cell culture chambers to allow filling and emptying of the cell culture chambers of liquid medium. 
     In an eighteenth aspect, there is provided the cell culture apparatus of aspect 17, wherein the column structure segments of adjacent manifold segments are interconnected by a sealing ring. 
     In a nineteenth aspect, there is provided the cell culture apparatus of aspect 18, wherein the sealing ring seals an interface between the column structure segments of adjacent manifold segments. 
     In a twentieth aspect, there is provided the cell culture apparatus of aspect 18, wherein each column structure segment comprises a shroud structure, wherein adjacent shroud structures face each other thereby forming a partial enclosure that extends about an entire periphery of the sealing ring. 
     In a twenty-first aspect, a cell culture apparatus, comprises: a cell culture modules comprising multiple cell culture chambers; a fluid manifold that connects the cell culture chambers together, the fluid manifold comprising a side wall base structure and a column structure, the column structure defining a fluid flow pathway to allow filling and emptying of the cell culture chambers; and an air manifold that connects the at least two cell culture manifolds together, the air manifold comprising a side wall base structure and a column structure, the column structure of the air manifold defining a fluid flow pathway to allow filling and emptying of the cell culture chambers; wherein the column structure of the air manifold comprises one or more indent structure that provides the column structure of the air manifold with a necked-down region. 
     According to a twenty-second aspect, there is provided the cell culture apparatus of aspect 21, wherein the air manifold comprises manifold segments, each manifold segment is associated with one of the at least two cell culture modules and each manifold segment comprises a side wall base structure segment and a column structure segment that is formed as a monolithic part of the side wall base structure segment. 
     According to a twenty-third aspect, there is provided the cell culture apparatus of aspect 22, wherein each column structure segment comprises an indent structure. 
     According to a twenty-fourth aspect, there is provided the cell culture apparatus of any of aspects 21-23, further comprising a riser having an internal volume that is located between the column structure of the air manifold and an air outlet tube of the column structure of the air manifold. 
     According to a twenty-fifth aspect, there is provided the cell culture apparatus of any of aspects 21-24, further comprising an air outlet tube that is in fluid communication with the column structure of the air manifold, the air outlet tube having a central axis that is offset laterally from a central axis of the column structure of the air manifold. 
     In an additional aspect, there is provided a cell culture apparatus, comprising a cell culture module comprising multiple cell culture chambers; each cell culture chamber having a top, a bottom and sidewalls, defining an interior space for culturing cells; wherein each cell culture chamber comprises at least one inlet in a sidewall of the cell culture chamber through which liquid can flow into and out of the cell culture chamber; wherein the multiple cell culture chambers are stacked one above the other to form the cell culture module; a manifold comprising a side wall base structure and a column; wherein the manifold is aligned along a side of the cell culture module; wherein the manifold provides a fluid pathway from a manifold opening to each of the cell culture chambers through cell culture inlets; wherein the manifold opening extends above the top-most stacked cell culture chamber; and wherein the column provides an enlarged volume inside the manifold to enable fluid to flow through the manifold to the cell culture chamber inlets. 
     Culture apparatuses described herein may be stacked into multi-layer culture chamber assemblies where individual chambers or groups of chambers are connected to each other via one or more manifolds. The manifolds include column structures that are formed as a monolithic part of the manifolds. The column structures may allow for use of the culture apparatuses as closed or open systems and may facilitate filling and/or emptying the culture apparatuses without any need for turning or repositioning the culture assemblies only during the filling and/or emptying processes. This and other advantages will be readily understood from the following detailed descriptions when read in conjunction with the accompanying drawings. 
    
    
     
       BRIEF DESCRIPTION OF THE DRAWINGS 
         FIG. 1  is a perspective view of a cell culture apparatus including manifolds, according to one or more embodiments shown and described herein; 
         FIG. 2  is a schematic view of a stack of stackette layers for use with the cell culture apparatus of  FIG. 1 , according to one or more embodiments shown and described herein; 
         FIG. 3A  is a schematic view of multiple module adapters for use with the cell culture apparatus of  FIG. 1 , according to one or more embodiments shown and described herein; 
         FIG. 3B  is a schematic view of a unitary manifold adapter for use with the cell culture apparatus of  FIG. 1 , according to one or more embodiments shown and described herein; 
         FIG. 3C  is a schematic view of a manifold including a column structure for use with the cell culture apparatus of  FIG. 1 , according to one or more embodiments shown and described herein; 
         FIG. 3D  is a side schematic view of the manifold including column structure of  FIG. 3   c;    
         FIG. 4  is a schematic view of another manifold for use with the cell culture; apparatus of  FIG. 1 , according to one or more embodiments shown and described herein; 
         FIG. 5  is a section view along lines  5 - 5  of the cell culture apparatus of  FIG. 1 , according to one or more embodiments shown and described herein; 
         FIG. 6  is a schematic view of another embodiment of a cell culture apparatus including a manifold, according to one or more embodiments shown and described herein; 
         FIG. 7  illustrates a filling operation where multiple cell culture apparatuses are filled in parallel, according to one or more embodiments shown and described herein; 
         FIG. 8  illustrates another filling operation where multiple cell culture apparatuses are filled is series, according to one or more embodiments shown and described herein; 
         FIG. 9  illustrates another embodiment of a cell culture apparatus including manifolds, according to one or more embodiments shown and described herein; 
         FIG. 10  illustrates a section view of the cell culture apparatus along lines  10 - 10  of  FIG. 9 , according to one or more embodiments shown and described herein; 
         FIG. 10A  illustrates a method of assembling the manifold of the cell culture apparatus of  FIG. 9 , according to one or more embodiments shown and described herein; 
         FIG. 11  is a schematic view of another embodiment of a manifold for use with a cell culture apparatus, according to one or more embodiments shown and described herein; 
         FIG. 12  illustrates a filling operation where multiple cell culture apparatuses are filled in parallel, according to one or more embodiments shown and described herein; 
         FIG. 13  illustrates another filling operation where multiple cell culture apparatuses are filled is series, according to one or more embodiments shown and described herein; 
         FIG. 14  illustrates another embodiment of a manifold for use with a cell culture apparatus, according to one or more embodiments shown and described herein; 
         FIG. 15  illustrates a front view of another embodiment of a cell culture apparatus, according to one or more embodiments shown and described herein; 
         FIG. 16  illustrates another front view of the cell culture apparatus of  FIG. 15 ; 
         FIG. 17  illustrates a side view of the cell culture apparatus of  FIG. 16 ; 
         FIG. 18  illustrates a section view of the cell culture apparatus of  FIG. 16 ; 
         FIG. 19  is a section view of the cell culture apparatus of  FIG. 15  along lines  19 - 19  illustrating a fill process, according to one or more embodiments shown and described herein; 
         FIG. 20A  is the section view of  FIG. 19  illustrating a fill line; 
         FIG. 20B  is the section view of  FIG. 19  illustrating the fill line progression; and 
         FIG. 20C  is the section view of  FIG. 19  illustrating the fill line progression. 
     
    
    
     The drawings are not necessarily to scale. Like numbers used in the figures refer to like components, steps and the like. However, it will be understood that the use of a number to refer to a component in a given figure is not intended to limit the component in another figure labeled with the same number. In addition, the use of different numbers to refer to components is not intended to indicate that the different numbered components cannot be the same or similar. 
     DETAILED DESCRIPTION 
     In the following detailed description, reference is made to the accompanying drawings that form a part hereof, and in which are shown by way of illustration several specific embodiments of devices, systems and methods. It is to be understood that other embodiments are contemplated and may be made without departing from the scope or spirit of the present disclosure. The following detailed description, therefore, is not to be taken in a limiting sense. 
     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. 
     As used in this specification and the appended claims, the singular forms “a”, “an”, and “the” encompass embodiments having plural referents, unless the content clearly dictates otherwise. As used in this specification and the appended claims, the term “or” is generally employed in its sense including “and/or” unless the content clearly dictates otherwise. 
     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”. 
     The present disclosure describes, inter alia, cell culture modules include a plurality of growth or culture surfaces in cell culture chambers coupled together via manifolds to form the cell culture modules. The cell culture modules can be further coupled to additional cell culture modules via manifolds to form stacked cell culture apparatuses. The plurality of culture surfaces may be stacked in a multi-layer configuration. The manifold includes an integral column structure that is formed as a monolithic part of the manifold. The column structure includes an inlet port and provides at least part of a fluid flow pathway from the inlet port that is in fluid communication with the individual cell culture chambers within the cell culture modules. The manifold may be configured to allow filling of the individual or groups of the cell culture chambers from the top down, bottom up and/or simultaneously depending, at least in part, on the manifold and column structure configuration. The manifolds and associated column structures may provide a closed system where the column structures can be connected to flexible tubing to isolate the cell culture chambers from the environment during use of the cell culture apparatuses. In other embodiments, the manifolds and associated column structures may provide an open system where fluids can be poured from the environment directly into the column structures (e.g., by removing a cap). 
     Nearly any cell culture article having a plurality of stacked layers or that can be stacked to form layers can be adapted to include a manifold as described herein. Examples of such cell culture articles include T-flasks, TRIPLE-FLASK cell culture vessels (Nunc., Intl.), HYPERFlask™ cell culture vessels (Corning, Inc.), CellSTACK™ culture chambers (Corning, Inc.), CellCube® modules (Corning, Inc.), CELL FACTORY culture apparatuses (Nunc, Intl.), HYPERStack™ and cell culture articles as described in U.S. Pat. No. 9,752,111, titled Cell Culture System with Manifold. 
     Referring to  FIG. 1 , a cell culture apparatus  10  includes three cell culture modules  12 ,  14  and  16 , each containing multiple cell culture chambers  18 , which are stacked, one on top of the other, to form the multiple layer cell culture apparatus  10 . Each cell culture chamber  18  is an individual cell culture layer having a top, a bottom and sidewalls which define an interior space for culturing cells. In embodiments, the top, bottom, sidewalls, or a combination of these may be gas permeable. In embodiments, there is an inlet in the sidewall of each cell culture chamber adjacent to or coupled to a manifold  20  and  22  to allow fluid to flow from the manifold  20  or  22  into each cell culture chamber  18 . Each cell culture module  12 ,  14  and  16  utilizes two manifolds  20  and  22 . Liquid may enter and exit the cell culture chambers of modules  12 ,  14  and  16  through the first manifold  20 . Thus, the first manifold  20  may be referred to as a fluid manifold. Air may enter and exit the cell culture modules  12 ,  14  and  16  through the second manifold  22 . Thus, the second manifold  22  may be referred to as an air manifold. 
     As shown in  FIG. 1 , the inlet, which may be a barbed inlet  34 , extends above the top-most stackette or cell culture chamber  18  when the device is arranged in cell culture configuration—so that the cell culture chambers  18  are horizontal. 
     The configuration of cell culture chambers  18  is shown in  FIG. 2 . The cell culture modules  12 ,  14  and  16  may each include multiple stackette layers,  24 , also called cell culture chambers  18  that, when stacked together, form the multiple cell culture chambers  18  having tracheal spaces (air spaces)  25  therebetween.  FIG. 2  is a schematic representation of the multiple stackette layers  24  that are stacked together to form the layered cell culture chambers  18 . The cell culture chambers  18  may each be defined by a generally transparent bottom cell culture surface  26  and a generally transparent top surface  29  and also by sidewalls (not shown). The cell culture surfaces  26  are adjacent to tracheal spaces  25 . The cell culture surfaces  26  may include a gas permeable, liquid impermeable film  28 . This allows for the transfer of gasses between the cell culture chambers  18  and the tracheal spaces  25  which are in gas communication with the exterior of the cell culture apparatus  10 , through the gas permeable, liquid impermeable film  28 . When cells are cultured in the cell culture chambers  18 , they fall by gravity onto the cell culture surfaces  26  where they are bathed by cell culture media in the cell culture chambers  18 , and they are able to access oxygen through the gas permeable, liquid impermeable film  28  upon which they reside. Referring back to  FIG. 1 , the cell culture modules  12 ,  14  and  16  may be separated from one another by spacers  31 ,  33  and  35 . The spacers  31 ,  33  and  35  can provide structural support for the individual cell culture modules  12 ,  14  and  16 . In some embodiments, spacers  31  and/or  33  may be replaced by additional stackette layers  24  to provide a higher total number of cell culture chambers  18 . Stackettes  24  which define cell culture chambers  18 , are stacked together to form modules  12 ,  14  and  16 . Further, a riser volume may be provided above the cell culture module  12  to catch residual air, rather than air residing in the cell culture chambers  18 . 
     A cell culture module, or portions thereof, as described herein may be formed from any suitable material. Preferably, materials intended to contact cells or culture media are compatible with the cells and the media. Typically, cell culture modules are formed from polymeric material. Examples of suitable polymeric materials include polystyrene, polymethylmethacrylate, polyvinyl chloride, polycarbonate, polysulfone, polystyrene copolymers, fluoropolymers, polyesters, polyamides, polystyrene butadiene copolymers, fully hydrogenated styrenic polymers, polycarbonate PDMS copolymers, and polyolefins such as polyethylene, polypropylene, polymethyl pentene, polypropylene copolymers and cyclic olefin copolymers, and the like. 
     In some embodiments, the culture units contain the gas permeable, liquid impermeable film  28  to allow transfer of gasses between the cell culture chamber  18  and ultimately with the exterior of the cell culture assembly. Such culture units can include spacers or spacer layers positioned adjacent the film, exterior to the chamber, to allow air flow between stacked units. One commercially available example of a cell culture apparatus containing such stacked gas permeable culture units is Corning&#39;s HYPERStack™ cell culture apparatus. Examples of suitable gas permeable polymeric materials useful for forming a film include polystyrene, polyethylene, polycarbonate, polyolefin, ethylene vinyl acetate, polymethylpentene, polypropylene, polytetrafluoroethylene (PTFE), or compatible fluoropolymer, a silicone rubber or copolymer, poly(styrene-butadiene-styrene) or combinations of these materials. As manufacturing and compatibility for the growth of cells permits, various polymeric materials may be utilized. Preferably the film is of a thickness that allows for efficient transfer of gas across the film. For example, a polystyrene film may be of a thickness of about 0.003 inches (about 75 micrometers), though various thicknesses are also permissive of cell growth. As such, the film may be of any thickness, preferably between about 25 and 250 micrometers, or between approximately 25 and 125 micrometers. The film allows for the free exchange of gases between the chamber of the assembly and the external environment and may take any size or shape. Preferably, the film is durable for manufacture, handling, and manipulation of the apparatus. 
     As mentioned above with reference to  FIG. 1 , the cell culture modules  12 ,  14  and  16  and their individual cell culture chambers  18  may be connected together using the manifolds  20  and  22 . That is, manifolds  20  and  22  provide a fluid pathway to introduce media and cells into the manifold, and into individual cell culture chambers  18 . The manifold  20  includes a side wall base structure  30  and a column structure  32  that is formed as a monolithic part of the side wall base structure  30  providing a unitary manifold  20 . The column structure provides an enlarged volume inside the manifold to enable fluid to flow through the manifold to the cell culture chambers  18 . Manifold includes a manifold opening, shown in  FIG. 1  as a barb structure  34 . In embodiments, the manifold opening extends above the top-most stacked cell culture chamber  18 , when the apparatus is arranged in cell culture orientation. The column structure  32  includes a barb structure  34  and provides at least part of a fluid flow pathway from the barb structure  34  to the individual cell culture chambers  18  within the cell culture modules  12 ,  14  and  16 . The manifold  20  may be configured to allow filling and emptying of the cell culture chambers  18  through, for example, ports  52  (as shown in  FIG. 2 ). The column structure  32  provides an enlarged volume to allow for faster and more efficient filling and emptying of the cell culture chambers  18 . 
     The manifold  22  also includes a side wall base structure  30 ′ and a column structure  32 ′ that is formed as a monolithic part of the side wall base structure  30 ′ providing a unitary manifold  22 . The column structure  32 ′ includes a barb structure  34 ′ and provides at least part of a fluid flow pathway from the individual cell culture chambers  18  within the cell culture modules  12 ,  14  and  16  to the barb structure  34 ′. The manifold  22  may be configured to allow filling and emptying of the cell culture chambers  18  by allowing air to enter and exit the cell culture apparatus  10 . 
     Referring to  FIGS. 3A-3C , an exploded, schematic view of an optional manifold assembly  36  including the manifold  20  is illustrated. Referring first to  FIG. 3A , a front view of module adapters  40 ,  42  and  44  are illustrated. The module adapters  40 ,  42  and  44  are formed of a suitable material that is used to seal against each of the cell culture modules  12 ,  14  and  16  individually and to provide a seal between the cell culture chambers  18  and a unitary manifold adapter  46 . As can be seen, the module adapters  40 ,  42  and  44  each include a perimeter  48  that seals to the manifold adapter  46 . The module adapters  40 ,  42  and  44  also include a module adapter body  50  with multiple ports  52  that each provides ingress and egress to the individual cell culture chambers  18 . In embodiments, when an apparatus having three modules is assembled, these module adapters may be affixed to the modules to allow for efficient connections between the manifold  20  and the modules  48 . 
       FIG. 3B  illustrates an optional unitary manifold adapter  46 . The unitary manifold adapter  46  is formed of a suitable material that is used to seal between all of the module adapters  40 ,  42  and  44  and the side wall base structure  30  of the manifold  20 . The unitary manifold adapter  46  includes a perimeter  48  and intermediate portions  49  and  51  that seal to the side base wall of the manifold  20 . The unitary manifold adapter  46  also include a unitary manifold adapter body  54  that seals to the perimeters  48  of the module adapters  40 ,  42  and  44 , thereby defining module openings  56 ,  58  and  60 . Each module opening  56   58  and  60  is sized to expose the ports  52  of the module adapters  40 ,  42  and  44  to the fluid flow path between the manifold  20  and the cell culture chambers  18 . 
     Referring to  FIG. 3C , the manifold  20  is illustrated and includes the side wall base structure  30  and the column structure  32  that is formed as a monolithic part of the side wall base structure  30  providing the unitary manifold. The side wall base structure  30  seals against the unitary manifold adapter  46  thereby forming a fluid-tight seal between the manifold  20  and the cell culture modules  12 ,  14  and  16 . In some embodiments, the column structure  32  may include the barb structure  34  that can facilitate connection with a tube for a filling or emptying operation in a closed system. The barb structure may be a component separate from the column structure  32 , or may be monolithic with the column structure  32 . If the barb structure  34  is a separate component, any suitable method of attaching the barb structure  34  to the column structure  32  may be used, such as adhesive, thermal bonding, ultrasonic welding, infrared welding, laser welding, etc. Separately forming the barb structure  34  can allow for increased access to the cell culture apparatus  10  during assembly. Forming the barb structure  34  monolithic with the column structure  32  can eliminate a step of attaching the barb structure  34  to the column structure  32 .  FIG. 3D  illustrates a side view of the manifold  20 . 
     Referring briefly to  FIG. 4 , the manifold  22  and associated manifold assembly  66  may include the same features as the manifold assembly  36  of  FIGS. 3A-3C . The side wall base structure  30 ′ seals against a unitary manifold adapter  36 ′ and module adapters  40 ′,  42 ′ and  44 ′ thereby forming a fluid-tight seal between the manifold  22  and the cell culture modules  12 ,  14  and  16 . 
     Referring to  FIG. 5 , a section view of the cell culture apparatus  10  and manifold assembly  36  are illustrated through line  5 - 5  of  FIG. 1 . The manifold assembly  36  includes the manifold  20  that includes the column structure  32 . The column structure  32  provides the fluid flow path P that extends between the manifold  20  and the module adapters  40 ,  42  and  44  and leads to the cell culture chambers  18  that are stacked one over the other, via the ports  52 . In particular, the unitary manifold adapter  46  is located behind the manifold  20  and seals between the manifold  20  and the module adapters  40 ,  42  and  44 . In some embodiments, an adhesive or other sealant material  68  may be used between the manifold  20 , unitary manifold adapter  36  and module adapters  40 ,  42  and  44 . Thus, the fluid flow path P extends from the barb structure  34 , through the column structure  32  and into the cell culture chambers  18  through the ports  52  that provide ingress and egress to the cell culture chambers  18 . The column structure  32  provides an enlarged volume to improve fluid flow into and out of the cell culture chambers  18 , which improves the speed at which the device can be filled and emptied. 
     The components can be joined by any suitable method including adhesive, thermal bonding, ultrasonic welding, infrared welding, laser welding, etc. To accommodate laser welding, a black color may be compounded into a shot of polymer that extends about a periphery of each adapter. 
       FIG. 6  illustrates a schematic view of a 12 layer (single cell culture module) cell culture apparatus  70  with a manifold assembly  72  including a module adapter  74 . In embodiments, the unitary manifold adapter described above may be omitted. 
     Referring to  FIG. 7 , multiple of the cell culture apparatuses  10  are illustrated during a filling operation. In this example, the cell culture apparatuses  10  are all connected at the column structures  32  to a common fluid inlet line  76  for filling in parallel. The column structures  32 ′ are connected to a filter  75   FIG. 8  illustrates multiple cell culture apparatuses  10  connected to each other using both the manifolds  20  and  22  for filling in series. As can be seen, a filling operation can be accomplished without any need for turning the cell culture apparatus  10  on its side and then turning the cell culture apparatus  10  upright, which can be heavy. Or, in embodiments, as shown for example in  FIG. 19 , the device can be turned on its side for filling, or elevated at a corner for filling. 
     Referring to  FIG. 9 , another embodiment of a cell culture apparatus  100  includes three cell culture modules  102 ,  104  and  106 , each containing multiple layers of cell culture chambers  108 , as described above. In this embodiment, the cell culture modules  102 ,  104  and  106  utilize a manifold  110  that is formed of multiple manifold segments  112 ,  114  and  116  that are connected together by column structure segments  118 ,  120  and  122  that are formed as monolithic parts of their respective manifold segments  112 ,  114  and  116 . As can be seen, the other air flow manifold is removed from the cell culture apparatus  100  to expose module adapters  124 ,  126  and  128 , which are similar or the same as those module adapters described above. 
     Referring also to  FIG. 10 , a section view of the cell culture apparatus  100  and manifold segments  112 ,  114  and  116  are illustrated through line  10 - 10  of  FIG. 9 . The manifold  110  includes the manifold segments  112 ,  114  and  116 , where each manifold segment  112 ,  114  and  116  is associated with a cell culture module  102 ,  104  and  106 . The manifold segments  112 ,  114  and  116  each include a side wall base structure  130 ,  132  and  134  and a column structure segment  118 ,  120  and  122  that is formed monolithically with the respective side wall base structure  118 ,  120  and  122 . 
     Referring particularly to  FIG. 10 , a section view of the cell culture apparatus  100  and manifold assembly  136  are illustrated through line  10 - 10  of  FIG. 9 . The manifold assembly  136  includes the manifold  110  that includes the manifold segments  112 ,  114  and  116 , each including the column structure segment  118 ,  120  and  122 . The column structure segments  118 ,  120  and  122  provide a fluid flow path P that leads to cell culture chambers  138  that are stacked one over the other, as described above. The module adapters  124 ,  126  and  128  are located behind the manifold segments  112 ,  114  and  116  and seal between the manifold segments  112 ,  114  and  116  and the cell culture modules  102 ,  104  and  106 . 
     The column structure segments  118 ,  120  and  122  are interconnected using sealing rings  140  and  142 . The sealing rings  140  and  142  may be received within shroud structures  146   a ,  146   b  and  148   a ,  148   b  and surround interfaces  152  and  154  between the column structure segments  118 ,  120  and  122 . The sealing rings  140  and  142  may be formed of any material suitable for both connecting the column structure segments  118 ,  120  and  122  together, but also to provide a sealing function, such as plastics or rubber. The sealing rings  140  and  142  may be releasably used or permanently affixed between shroud structures  146  the column structure segments  118 ,  120  and  122  such as through welding, adhesives, overmolding, etc. Further, the sealing rings  140  and  142  may be formed of a relatively stiff or flexible materials, or combinations thereof. The sealing rings  140 ,  142  may be formed of an elastomer and include thicker portions  151  and  153  that optionally are separated by a relatively thinned portion  155  (e.g., a notch) that defines an area of increased flexibility that can facilitate sealing against the column structure segments  118 ,  120  and  122  at the interfaces  152 . The thicker portions  151  and  153  have a lead in taper at both ends  157  and  159  to enable for ease of assembly. 
       FIG. 10A  illustrates a method of assembling the column structure segments  118 ,  120  and  122  together with the sealing rings  140  and  142 . The column structure segment  118  is an upper column structure segment that includes an upper end  182  that can be formed as a barb structure  184  and a lower end  186  that is received within the sealing ring  140 . In some embodiments, the lower end  186  tapers in wall thickness to facilitate insertion of the lower end  186  into the sealing ring  140 . The column structure segment  120  is an intermediate column structure segment that includes an upper end  188  that is received within the sealing ring  140  and a lower end  190  that is received within the sealing ring  142 . The lower end  190  and the upper end  188  may taper in wall thickness to facilitate insertion of the upper end  188  into the sealing ring  140  and the lower end  190  into the sealing ring  142 . The column structure segment  122  is a lower column structure segment that includes an upper end  192  that is received within the sealing ring  142  and a lower end  194  that is closed to prevent passage of fluid therethrough. The upper end  192  may taper in wall thickness to facilitate insertion of the upper end  192  into the sealing ring  142 . 
     Each column structure segment  118 ,  120  and  122  includes the shroud structure  146   a ,  146   b ,  148   a ,  148   b . As assembled, adjacent shroud structures  146   a ,  146   b  and  148   a  and  148   b  face one another forming a partial enclosure that extends about the entire periphery of the sealing rings  140  and  142 . This partial enclosure that is formed by the adjacent shroud structures  146  can protect the sealing rings  140  from contact and maintain seal integrity. 
     In embodiments, the modular cell culture device may be assembled by stacking one cell culture module on another cell culture module, each cell culture modules comprising multiple cell culture chambers; and connecting the cell culture chambers using a manifold, the manifold comprising a side wall base structure and a column that is formed as a monolithic part of the side wall base structure, the column structure defining an enlarged fluid flow pathway through the manifold and to the cell culture chambers to allow filling and emptying of the cell culture chambers of liquid medium. 
     Referring to  FIG. 11 , another embodiment of a manifold  160  is illustrated for use with a manifold assembly  165  that is similar to the manifold of  FIG. 3 c   . In this embodiment, an inlet opening  162  of column structure  164  that is a monolithic part of the manifold  160  is located at a bottom  166  of the manifold  160 , as opposed to the top. The column structure  164  may include a bend  168  to that the inlet opening  162  faces outward to facilitate filling and emptying operations. 
       FIGS. 12 and 13  illustrate parallel and series filling, respectively, of multiple cell culture apparatuses  165  using the manifold of  FIG. 11 . Filling may occur when the devices are upright, as shown in  FIG. 12 , or on their sides, as shown, for example, in  FIG. 19 . 
     Referring to  FIG. 14 , another embodiment of a manifold  170  for use with a manifold assembly  172  includes a column structure  174  that is a monolithic part of the manifold  170 . In this embodiment, the column structure  174  has an inlet opening  176  that is openable and closable using a closing structure  178 , such as a cap. Such a column structure arrangement can provide an open system where the cell culture apparatus can be filled by pouring fluid into the column structure  174 . The manifold can be used for one or multiple cell culture modules. 
     Referring to  FIGS. 15-17 , another embodiment of a cell culture apparatus  200  includes many of the features provided by the cell culture apparatus  100  of  FIG. 9  including three cell culture modules  202 ,  204  and  206 , each containing multiple layers of cell culture chambers  208 . As with the cell culture apparatus of  FIG. 9 , the cell culture modules  202 ,  204  and  206  utilize a manifold  210  that is formed of multiple manifold segments  212 ,  214  and  216  that are connected together by column structure segments  218 ,  221  and  222  that are formed as monolithic parts of their respective manifold segments  212 ,  214  and  216 . Another manifold  220  is an air flow manifold that is also formed of multiple manifold segments  222 ,  224  and  226  that are connected together by column structure segments  228 ,  230  and  232  that are formed as monolithic parts of their respective manifold segments  222 ,  224  and  226 . 
       FIG. 15  illustrates inlet and outlet tubes  234  and  236  that are connected to barb structures  238  and  240  ( FIG. 16 ) that provide fluid communication with the column structure segments  218 ,  221 ,  222  and  228 ,  230 ,  232 , respectively. Each inlet and outlet tube  234  and  236  is connected to its respective barb structure  238  and  240  by a connector  242 . The connectors  242  may have a locking connection with the barb structure  238  and  240  to inhibit unintended removal of the inlet and outlet tubes  234  and  236  from the barb structures  238  and  240  and also to promote a sealing engagement between the connectors  242  and the barb structures  238  and  240 . 
     Referring also to  FIG. 18 , the manifolds  210  and  220  are provided with risers  244  and  246  having internal volumes  248  and  250  that are in fluid communication with column structure segments  218  and  228 . The risers  244  and  246  provide the internal volumes  248  and  250  located at least partially above the cell culture module  202  to provide a holding location for liquid and/or air during a filling operation. The riser  246 , in particular, provides the internal volume  250  that is sized to allow air to escape the column structure segments  228 ,  230  and  232  during the filling operation, which facilitates filling of the cell culture modules  202 ,  204  and  206  and reduces pockets of air from being trapped within the manifolds  210  and  220 . Providing the riser  246  and internal volume  250  can also allow for a reduction in volume of the column structure segments  228 ,  230  and  232 , which can also facilitate evacuation of air during the filling operation and reduce pockets of air from being trapped within the manifolds  210  and  220 . 
     In particular, each column structure segment  228 ,  230  and  232  includes an indent structure  254  that provides each column structure segment  228 ,  230  and  232  with a necked-down region providing a reduced volume compared to the absence of the indent structures  254 . The indent structures  254  also provide the column structure segments  228 ,  230  and  232  with volumes that are different than the volumes of the column structure segments  218 ,  220  and  222 , thereby providing an asymmetric column structure volume arrangement between column structures  250  and  252 . In some embodiments, the internal volume  250  of the riser  246  may be selected based on the volume reduction provided by the indent structures  254 . In some embodiments, the internal volume  250  of the riser  246  may be at least 50 percent of the volume reduction provided by the indent structures  254 , such as at least 75 percent, such as at least 100 percent or more. 
     Referring particularly to  FIG. 18 , another difference between the cell culture apparatus  200  and the cell culture apparatus  100  of  FIG. 9  is that the barb structure  240  provides an air outlet  260  that is offset from the column structure  252 . In particular, the air outlet  260  has a central axis A 1  that is offset laterally from a central axis A 2  of the column structure  252  provided by the column structure segments  228 ,  230  and  232 . The offset arrangement of the air outlet  260  inhibits the flow of liquid medium into the air outlet  260 , which can then reach an air filter connected to the air outlet  260 . 
     Referring to  FIG. 19 , during a fill operation, the cell culture apparatus  200  may be tilted and supported on a spacer block  300 . The cell culture apparatus  200  is then filled through the manifold  210  and air is evacuated through the manifold  220 . 
       FIGS. 20A-20C  illustrate the filling progression approaching the end of the filling process showing fill line F. The shapes of the indent structures  154  and/or the riser  246  can be selected to reduce or even eliminate air pockets within the manifold  220  once the entire cell culture apparatus  200  is filled. As can be seen by  FIG. 20C , the shape of the indent structure  154  may be selected to provide a wall that is shaped to be flush with the fill line F with the cell culture apparatus  200  tilted to a predetermined angle provided by the spacer block  300 . Additionally, the offset arrangement of the air outlet  260  allows for complete filling of the manifold  220  with medium while reducing flow of liquid medium into the air outlet  260  as the air outlet is out of alignment with the column structure  252 . In addition, this angle of the indent structures  154  and/or the riser  246  allows for as nearly complete filling as possible, without over-filling. Over-filling of the device may cause media to spill out of the device as the device is manipulated and may cause contamination. For example, often a vent filter is present in the tubing  301  which extends from the device (see, for example,  FIG. 18 ). If the vent filter is wetted with protein-rich media, the vent filter may become clogged. A clogged vent filter may prevent the system from properly filling or emptying because the flow path is blocked. This may create a contamination risk if a user removes the vent filter either to wash it or to allow the device to flow. Therefore, providing the indent structure  154  and the riser angles allows for more complete filling of the device without over-filling which can result in contamination or obstruction of the fluid flow path. 
     Manifolds, or portions thereof, as described herein may be formed from any suitable material. For example, a manifold, or component thereof may be formed from a biocompatible polymeric material. In various embodiments, a manifold is formed from one or more materials from which a cell culture module is formed. 
     It will be understood that a manifold or cell culture module may be of any suitable size. In many of the depicted embodiments, the column structures or components thereof are depicted as having a rounded cross-sectional shape, but it will be understood that they may have any suitable cross-sectional shape, such as rectangular, ellipsoidal or the like. It will be further understood that a cell culture module may include any number of cell culture chambers. In some embodiments, a cell culture module has, for example, 10 stacked cell culture chambers or 12 stacked cell culture chambers. Multiple ones of the cell culture modules may be stacked, one on the other, to form a cell culture apparatus. In embodiments, any number of stacked cell culture chambers may be assembled in cell culture modules, providing manifolds with any number of cell culture chambers. 
     Thus, embodiments of CELL CULTURE APPARATUSES WITH MANIFOLDS INCLUDING COLUMN STRUCTURES are disclosed. One skilled in the art will appreciate that the cell culture apparatuses and methods described herein can be practiced with embodiments other than those disclosed. The disclosed embodiments are presented for purposes of illustration and not limitation.