Deformable transportable bioreactor chamber

An apparatus and method is described for seeding and culturing cells on a sample. The apparatus includes a chamber in which the volume of the chamber may be adjusted without compromising the seal or sterility of the chamber. The apparatus enables the seeding of cells in a reduced volume and culturing of cells in an increased volume. Further, the apparatus enables application of forces, strains and torques to a sample during seeding, culturing or transportation of the sample.

CROSS-REFERENCE TO RELATED APPLICATIONS

Not Applicable

FEDERAL SPONSORSHIP

Not Applicable

JOINT RESEARCH AGREEMENT

Not Applicable

TECHNICAL FIELD

This invention pertains generally to cell seeding and cell tissue growth. This invention also pertains to flexible, deformable, chambers suitable for seeding and growing cells on a sample within a sterile interior of the chamber.

BACKGROUND

Generally, the seeding or depositing of cells and subsequent growth or culture of cells has previously been described. In the past, cells have been seeded and cultured on a matrix, specimen, tissue, vascular graft, biomedical prosthesis, substrate, and other medical devices (hereinafter referred to simply as a sample or specimen). Some prior systems seed cells on a sample in a seeding chamber and then transfer the sample to a growth chamber, where nutrients are supplied to the cells for growth. Other systems have used pressure or other fluid forces to influence adhesion of the cells on the sample. It has been recognized that cells seeded or cultured in a dynamic fluidic environment are more likely to tolerate physiological conditions of the human body.

Other prior devices have cultured a sample within a disposable bag. It has been recognized that lack of a framework for the bag during transport is not preferred. Other user criteria may further influence the acceptance and use of a particular chamber including the ease of transport, scalability of the chamber to accommodate varying length and widths of a sample, and versatility of chamber for use while seeding, culturing or testing a sample in a sterile environment. Further, it is now recognized that it is advantageous to provide a dynamic environment that allows a constant or varying strain or other forces applied to the sample during seeding, culture, testing and transport.

SUMMARY

Embodiments of the invention include an apparatus and method for seeding, culturing, testing, and transporting a sample without removing the sample from a sterile chamber of the apparatus. The chamber has a volume that is adjustable such that cells may be seeded on the sample in a reduced volume and cells may be cultured on the sample in an expanded volume. Further, in an embodiment of the invention, the chamber includes a first deformable outer sheath and second deformable inner enclosure. Also, a dual membrane chamber of the invention may be transported while maintaining the sterility of the interior membrane of the chamber.

During seeding and culturing cells on the sample, linear forces, strains, and torques may be applied to the sample within the chamber. Further, the chamber may be transported while maintaining a linear force, strain or torque applied to the sample. Alternatively, in an embodiment of the invention, the volume of the chamber may be varied while applying varied linear forces, strains and torques on the sample. Also, in an embodiment of the invention the volume is varied with the aid of a mold that constricts a portion of the chamber, thereby reducing the volume. Additionally, fluids may be delivered into the chamber causing the chamber to expand to an increased volume.

Also described herein are grips contained within the chamber. The grips have an adjustable separation distance between the grips. Further, an installation and indexing frame is coupled to the chamber to facilitate a repeatable gauge length of multiple samples and to provide support to the chamber during transport. To facilitate loading of a sample in the grips a first end of the chamber may be drawn towards the second end to expose a space between the grips. In this manner the space between the grips is accessible from multiple angles.

The accompanying drawings, which are incorporated in and constitute a portion of this specification, illustrate embodiments of the invention and, together with the detailed description, serve to further explain the invention. The embodiments illustrated herein are presently preferred; however, it should be understood, that the invention is not limited to the precise arrangements and instrumentalities shown. For a fuller understanding of the nature and advantages of the invention, reference should be made to the detailed description in conjunction with the accompanying drawings.

DETAILED DESCRIPTION

The following description provides detail of various embodiments of the invention, one or more examples of which are set forth below. Each of these embodiments are provided by way of explanation of the invention, and not intended to be a limitation of the invention. Further, those skilled in the art will appreciate that various modifications and variations may be made in the present invention without departing from the scope or spirit of the invention. By way of example, those skilled in the art will recognize that features illustrated or described as part of one embodiment, may be used in another embodiment to yield a still further embodiment. Thus, it is intended that the present invention also cover such modifications and variations that come within the scope of the appended claims and their equivalents.

The bioreactor of the present invention includes a chamber capable of retaining fluids within the chamber and has a first expanded volume and second constricted volume. The volume of the chamber increases and decreases without adding or eliminating wall segments to the chamber. Alternatively, the chamber may include a first deformable outer sheath and second deformable inner enclosure. Ports may be coupled to the chamber and, in particular, may be coupled in fluid communication to the outer sheath and inner enclosure. Further, valves may be coupled to the ports to control the flow of fluids through the ports and may be utilized to assist hydrostatic pressure within the chamber.

The bioreactor includes grips containable within the chamber. A first end of each grip contained within the chamber is suitable for gripping a portion of a sample. A second end of each grip is suitable for coupling to a connector of the bioreactor. When coupled to the bioreactor, stepper motors or drivers of known suitable construction selectively deliver axial, linear and torsion loads on the grips while contained in the chamber. A controller may be utilized to control the motors and drivers and selectively apply forces, strains and torques to one or both grips. The chamber has first and second ends, wherein the first end may be drawn towards the second end to expose a space between the grips. With the space exposed, a sample is easily loaded and unloaded to and from the grips.

A user may use the apparatus to seed and culture cells on a sample within the chamber. The user positions a sample within the chamber. The sample may be held in place within the chamber by grips that are also contained within the chamber. A distance between the grips is controllable such that the gauge length of the sample held between the grips may be approximately equivalent among several samples. The chamber is capable of retaining fluids therein. Further, the user may selectively constrict the chamber so that it has a reduced volume during seeding and the chamber may be selectively expanded to have an increased volume during culturing and testing the sample. Those skilled in the art will appreciate that it may be desired to vary the volume of the chamber during the culturing and testing of the sample.

Once the sample is grasped between the grips, the user may selectively control the bioreactor such that linear forces, strains and torques are applied to the sample. This stimulus to the sample may be applied while the sample is contained within the chamber under a seal and may be applied before, during or after seeding and/or culturing the sample. The user may couple an installation and indexing frame to ends of the grip extending from the chamber. Once coupled to the grips, the chamber may be removed from the bioreactor. The indexing frame couples to the grips and retains the separation distance between the grips. Any torque, and axial or linear forces applied to the grips will also be retained in the indexing frame. In this manner, the user may transport the sample within the sterile chamber interior with a continuous force, strain or torque applied to the sample.

Turning attention now to the Figures, embodiments of the bioreactor or system10of the present invention will now be described in more detail. Referring first toFIGS. 1-2, the bioreactor10includes a frame100, drive assembly102, load cell104, quick disconnect coupling106, controller110, and mold cavity116supported by mold arms118. Although the chamber is shown inFIG. 2as opaque, those skilled in the art will appreciate that at least a portion of the chamber may be constructed from a translucent material. Drive assembly102may be coupled to a linear or axial actuator (not shown) contained with frame100. The drive assembly102and linear actuator may be of a servo pneumatic, electromechanical flexure bearing or electromechanical linear screw motors of known suitable construction. The drive assembly102and linear actuator are coupled to the load cell104, couplings106and frame100so that the couplings106may be rotated and the space between the couplings106may be increased or decreased in a controlled finite manner.

The controller110is electrically coupled to the drive assembly102, load cell104, linear actuator and sensors (not shown) so that feedback and analysis loops may be incorporated into the controller110to selectively provide repetitive, continuous, and intermittent stimulus to a sample60held in place between couplings106. As the connector106is rotated, a resulting torque is applied to the sample60. Further, the controller110may be utilized to alter a separation distance between the connectors106, thereby applying strains or axial and linear forces on the sample60. The controller110further allows the user to maintain the position of the connectors106in a fixed position to thereby translate a fixed strain, force or torque on the sample60. In this manner a variety of stimulus sequences may be applied to a selected sample60.

It will be appreciated by those skilled in the art that setting, monitoring and controlling the separation distance between the connectors enables finite control of the stimulus applied to the sample. Further, sensors may be electrically coupled to the controller110to detect the position of the top and bottom grips120. This displacement may be measured with, by way of illustration and without limitation, an LVDT, laser PSD, incremental encoder, or other measurement feedback device of known suitable construction. Under load control, the controller110adjusts the separation distance and positions of the couplings106and grips120so that a known force (common preload) may be applied to all samples60. The load cell104may also be utilized to control the force applied to the sample. Also, controlling the resulting distance between the grips120with a preload applied to the sample60allows the user to set a consistent gauge length to multiple samples.

FIG. 3shows a single chamber20of the present invention including a sheath or bag200having opposite ends affixed to feed through connectors108. Grips120engage and slide within the feed through connectors108and an o-ring or seal (refer toFIG. 11) restricts fluids from passing between the grip120and connector108. Alternatively, the seal may act as a deformable feed through that facilitates axial and/or rotational motion with respect to the connector108. For example, without limitation, the seal could include a mini bellow of known suitable construction. An end of each grip120is thereby sealed and contained within the sheath200. The other end of the grip120is coupled within a quick disconnect coupling106. The sample may be secured to the grips in a manner suitable to keep the ends of the sample from slipping in the grips. By way of example, grips120may be a suitable construction adapted for holding a tissue, vascular grafts, biomedical prosthesis, medical devices or other desired specimen or sample. Further, by way of example and without limitation intended, a tubular sample may slip over and be secured to an end of grips120and a relatively planar sample may be clamped or sutured to an end of the grips120.

Connectors108include a flange124formed on the end thereof and an end of the bag200is affixed to a shoulder126of the connector108. The flange124may serve as a stop for the end of bag200. Ports112, of suitable known construction, may be coupled to an exterior of the sheath200in a known manner to provide a seal between the port112and the sheath200. Further valves114may be coupled to the ports to control the ingress and egress of fluids through port112and to control hydrostatic pressure within the chamber. Further, ports112may be located at the top of the sheath200to facilitate delivery of cells and media into the interior of the chamber20and ports112may be located at the bottom of the sheath200to act as a drain and facilitate media exchange as well as acquiring media samples.

The exterior of each bag may include indicia, tags, chips or other device to identify the chamber20for tracking and monitoring the particular sample60contained within the chamber20. The sheath200is flexible and may include horizontal folds202and vertical folds (not shown). In an embodiment of the invention the various components of the chamber are manufactured from materials suitable for sterilization and preferably suitable for autoclave. The walls of sheath200,300and enclosure302may be constructed from a gas permeable, flexible, stable, durable, low durometer material capable to withstand autoclave, exposure to ethylene oxide, or gamma sterilization. Further, the preferably selected material is a translucent material that aids in the monitoring of the sample within the chamber. One such suitable material is a platinum cured silicone.

The sheath200,300and enclosure302may, for example, be constructed from two sheets of material welded together at seems, in accordance with techniques known in the art (for example, IV fluid bags). Alternatively, the sheath200,300and enclosure302may be molded, or extruded. Further, the ends of the sheath200,300and enclosure302may be sealed to the connector by heat, ultrasound or radiowave welding, or by other known suitable means of sealing them together. Alternatively, one end may be molded into the connector108.

FIGS. 4 and 5show generally a chamber30having a dual membrane. The chamber30includes an outer sheath300and inner enclosure302. Disc306is positioned between sheath300and enclosure302, separating an upper portion of the sheath300and enclosure302. A shoulder304of the disc306is sized to slip over the outside of connector108with an end of the inner enclosure302sandwiched and creating a seal between the disc306and connector108. An end of the sheath300is affixed to an outer surface of the shoulder304of disc306creating a seal between the sheath300and disc306. A groove308is formed on an outer edge of the disc306. A knife314blade follows the groove308to open the outer sheath300without compromising the sterile environment of the enclosure302(seeFIG. 12). Folds in the enclosure302and folds in the outer sheath300(not shown) may expand and open to increase a volume within the sheath300and enclosure302.

FIGS. 6 and 7show an index frame500coupled to a portion of the chamber20and bioreactor10. The frame500includes an expandable handle502having an expansion joint518, a base504, grabbers or fingers508. A v-groove510is formed in the ends of frame500and positively engage with an ends of the grips120. Finger508engages the grip on the opposite side and holds the grip120in the v-groove510. The base504is actuated by a spring506, and when squeezed towards the handle502an end of the finger508is pushed away from the v-groove510. Finger plate512keeps the finger508in place along the frame500. Pins514of the base504align with apertures516in the handle502and contain the springs506between the handle502and base504. An end of the frame500may also engage with the flange124on the connector108to provide a controlled separation distance between the connectors108. The expansion joint518may be adjusted to accommodate the length of the sample60, independent of whether or not the sample60is under preload conditions. The mechanism to engage or fasten index frame500to connectors108is not limited to a spring loaded clamping mechanism. Without limitation, other suitable mechanisms may be incorporated, including a thumb screw, set screws, magnets, electro magnets, adhesives, Velcro™, suction, or other known fasteners.

Having described the constructional features of embodiments of the invention, the mode of use will next be described. For discussion purposes, but without any limitation intended, use of the single chamber20will be described. With reference toFIG. 8, the user selects a chamber20and inserts each outer end of the grip120into the respective quick disconnect coupling106of bioreactor10. The couplings106are locked and engaged to the grip120so that a rotation of the coupling106will simultaneously rotate the grip120. The user then collapses the sheath200by drawing an upper end of the sheath200down towards a lower end of the sheath200exposing the interior ends of the grips20. The user may then engage a sample60in the grip ends of the grip120. The controller110is used to selectively determine the preload force and separation distance between the grips120. The user then draws open end208of the sheath200over the connector108and affixes the sheath to the connector in a known manner to create a seal between the sheath200and connector108.

Referring now toFIGS. 9-11, the bioreactor10is shown having the mold40articulated into position, such that the mold cavity constricts a portion of the chamber20. A portion210of the sheath200overlaps upper and lower ends of the mold40. The sheath200conforms to the shape of the cavity as illustrated inFIG. 11. In an embodiment not shown, the mold and mold cavity could be sized to encompass the entire chamber20within the mold cavity, whereby the entire sheath200would conform to the shape of the cavity. During seeding, a high concentration of cells in a small volume of media is delivered into the chamber through port112. After a desired lapse of time, the mold40is articulated away from the chamber and culture media or nutrients are introduced through the ports112to fill the sheath200and increase the volume of the chamber20. The volume of the chamber20may be adjusted by using the mold to partially constrict the sheath200.

Those skilled in the art will further appreciate that an expandable outer sheath or membrane300may be used to constrict an inner enclosure or membrane302. Once the cells have been cultured and the desired stimulus has been delivered to the sample, the user may then remove the chamber from the bioreactor10. The user may, for example, use the indexing frame500to grasp the chamber and remove the connectors108from the quick disconnect couplings106. If a strain was being applied to the sample at the time the chamber is removed, the frame may be adjusted so that the separation distance between the grips120remain constant, thereby keeping a strain applied to the sample60.

When transporting the chamber to and from a sterile environment, use of the dual membrane chamber30allows the user to transport the chamber in a non-sterile environment while maintaining the sterility of the inner membrane302. Once transported, the outer membrane300may be cut away without affecting the sterility of the inner membrane302. Likewise, in this manner, the sample may be seeded, cultured, and transported within the inner membrane302while maintaining the sterility of the sample.

These and various other aspects and features of the invention are described with the intent to be illustrative, and not restrictive. This invention has been described herein with detail in order to comply with the patent statutes and to provide those skilled in the art with information needed to apply the novel principles and to construct and use such specialized components as are required. It is to be understood, however, that the invention can be carried out by specifically different constructions, and that various modifications, both as to the construction and operating procedures, can be accomplished without departing from the scope of the invention. Further, in the appended claims, the transitional terms comprising and including are used in the open ended sense in that elements in addition to those enumerated may also be present. Other examples will be apparent to those of skill in the art upon reviewing this document.