WO2021206555A1

WO2021206555A1 - Fluidic device, cell culturing system and method of testing a compound

Info

Publication number: WO2021206555A1
Application number: PCT/NL2021/050231
Authority: WO
Inventors: Aisen Gabriel DE SÁ VIVAS; Andries Dirk VAN DER MEER
Original assignee: Universiteit Twente
Priority date: 2020-04-09
Filing date: 2021-04-09
Publication date: 2021-10-14
Also published as: NL2025320B1

Abstract

A fluidic device, in particular a microfluidic chip, for culturing a 3D cell culture comprises a bottom wall defining a bottom side of an open-top chamber arranged for containing a cell culture medium. A fluidic channel extends below the chamber between a channel inlet and a channel outlet for guiding a fluid flow. The bottom wall is provided with a micro-opening through which the chamber and the microfluidic channel are in fluid communication. A side wall extends from the bottom wall circumferentially around the chamber to a top wall opposite the bottom wall. An upper edge of the side wall delimits an open access upper side of the chamber and the top wall is arranged for closing the chamber at the upper side thereof and to be supported by the upper edge of the side wall. A sealing means is provided between opposing surfaces of the upper edge of the side wall and the top wall and arranged for reversible fluid sealing of the side wall and top wall when the chamber is closed at the upper side by the top wall.

Description

Title: Fluidic device, cell culturing system and method of testing a compound

Technical field

The invention relates to a fluidic device, in particular a microfluidic chip, for culturing a 3D cell culture comprising a bottom wall defining a bottom side of an open-top chamber arranged for containing a culture medium wherein a fluidic channel extends below the chamber between a channel inlet and a channel outlet for guiding a fluid flow, wherein the bottom wall is provided with a micro-opening through which the chamber and the microfluidic channel are in fluid communication, the fluidic device further comprising a side wall extending from the bottom wall circumferentially around the chamber to a top wall opposite the bottom wall, an upper edge of the side wall delimiting an open access upper side of the chamber; and the top wall arranged for closing the chamber at the upper side thereof, the top wall being supported by the upper edge of the side wall. The invention moreover relates to a cell culturing system comprising such a fluidic device. The invention also relates to a method of testing a compound, particularly a drug, on a cell culture, particularly a 3D cell culture, using such a fluidic device.

Background of the invention

A good understanding of uptake of biomolecules by cells under circumstances mimicking in vivo conditions may lead to improved knowledge of the effect of such biomolecules on a cellular, tissue, organ and subject level.

Particularly for the development of drug compounds and medicine, there is an increasing interest in means and methods that allow for investigation of cellular behavior of individual cells or larger cell cultures under in vivo conditions. Currently pharmacokinetic and toxicological evaluation of drug candidates relies largely on costly, labor-intensive, time-consuming and ethically questionable animal test systems, which show only very limited predictive value for clinical efficacy and toxicity. To reduce animal tests in new drug development, fluidic devices are more and more applied as relatively simple but effective tools which allow in vitro cell experimentation under controlled fluidic conditions mimicking fluidic conditions experienced by cells in vivo. Fluidic devices, and particularly microfluidic devices, have the necessary compartments such as cavities and channels to allow for three-dimensional cell culture and supply of fluids carrying e.g. nutrients, test pharmaceuticals and/or other cell culture compounds, while a behavior of the supplied fluids and an effect thereof on the cultured cells is highly predictable offering spatiotemporal control of low volumes and amounts of fluids and compounds being supplied to the cells. Fluidic devices allow for investigation, and control, of mechanical cues such as flow, perfusion, pressure, and mechanical stress, real-time imaging, and biochemical analysis in live cells. The small dimensions of spatially separated microfluidic compartments in fluidic cell culture devices allow of a multitude of individually controllable cell cultures in chambers on a single device. This facilitates high parallelization of experiments, high throughput of samples and reactions and thus improvement of reproducibility, as well as a reduction in reagent costs.

Particularly, fluidic devices allow for in vitro cell co-culture with cells constituting a specific tissue cultured in communication with cells constituting another tissue to represent multicellular organs or tissue-tissue interactions. For instance fluidic devices are employed to integrate multiple organ/tissue mimetics with active vascular conduits and barrier tissues.

Presently available microfluidic devices for in vitro 3D cell culture experimentation mostly comprise a closed system to shield the cell culture and the culture conditions from possible outside influences. This provides a limited accessibility to the cell culture grown in the culture chamber of the device. Thus the known devices render the simultaneous manipulation and analysis of cultured cells rather difficult, particularly for monitoring of cells in complex geometries with high spatial and temporal resolution and their individual retrieval during or following experiments.

As a result, there is a need for an improved microfluidic system for cell culture investigation which may be particularly applied in drug studies, vaccine development and other types of medical research. The invention thus has as an object to provide a new fluidic system, fluidic device and method for cell culture investigation, with which the above mentioned drawbacks are overcome. The invention has as a particular object to provide a new fluidic system, fluidic device and method for cell culture investigation with which it is possible to investigate all types of cells such as vascular cells and organ cells individually or in functional units of tissues and organs in vitro. These and other aspects of the invention are evident from the specification and claims hereinafter.

Summ ry of the invention

The fluidic device described herein addresses the above-discussed challenges and restrictions. In addition, the fluidic device described herein addresses additional challenges and restrictions associated with conventional fluidic devices, some of which are described below.

In one aspect a fluidic device for in vitro complex living tissue reconstruction is provided which closely mimics the in vivo tissue of a living multicellular organism such as a plant or animal.

In a particular aspect a fluidic device is provided in which two different cell systems in one device can be cultured, particularly a vascular system, e.g. a blood vessel, together with an organ system, e.g. heart cell tissue.

The invention provides a fluidic device for in vitro 3D cell culture experimentation comprising a bottom wall defining a bottom side of an open-top chamber arranged for containing a culture medium wherein a fluidic channel extends below the chamber between a channel inlet and a channel outlet for guiding a fluid flow, wherein the bottom wall is provided with an opening through which the chamber and the fluidic channel are in fluid communication, the fluidic device further comprising a side wall extending from the bottom wall circumferentially around the chamber to a top wall opposite the bottom wall, an upper edge of the side wall delimiting an open access upper side of the chamber; and the top wall arranged for closing the chamber at the upper side thereof, the top wall being supported by the upper edge of the side wall, wherein between opposing surfaces of the upper edge of the side wall and the top wall a sealing means is provided which sealing means is arranged for reversible fluid sealing of the side wall and top wall when the culture chamber is closed at the upper side by the top wall.

In the fluidic device of the invention a first cell culture, e.g. organ tissue cells such as cardiac cells, can be provided, maintained and/or cultured in a space in the open-top chamber to form an organ tissue cell structure, e.g. a heart or heart-mimicking 3D tissue structure. A second cell culture, e.g. vascular cells such as endothelial cells, can be provided, maintained and/or cultured in the fluidic channel to form a vascular tissue cell structure, e.g. a blood vessel and/or a capillary bed. The opening between the fluidic channel and open-top chamber allows for fusion of the cell cultures such that perfusion, diffusion and/or other exchange of compounds between the cell structures is enabled. Fluids can be supplied to the cell cultures via the inlet opening of the fluidic channel and the open access side of the open-top chamber. As such the cell tissues can be simultaneously supplied with different media and from different sides, e.g. inner and outer sides of the formed cell tissue. The open-top chamber further enables direct access to cells maintained in the chamber for easy and convenient seeding or retrieval of cells and/or stimulation of the cells.

To shield the cell cultures and the culture conditions from possible outside influences the open access side of the open- top chamber can, in use, i.e. during culturing, be covered by providing the top wall over the upper edge of the side wall such that the open access side is closed. In accordance with the invention the fluidic device has sealing means between opposing surfaces of the upper edge of the side wall and the top wall for reversible fluid sealing of the side wall and top wall when the open-top chamber is closed by the top wall. The sealing means enable the top wall of the fluidic device to be formed as a relatively simple lid or cover that is manually replaceable over the upper edge of the side wall as desired between an open state and a covered state of the open-top chamber. The cover or lid may be an integral part of the fluidic device or may be a separate body. Because of the sealing means a clamping force is not necessary to secure the top wall against the upper edge of the side wall, thereby obviating the need for mechanical clamping means. This importantly prevents cell-damaging pressure changes in the chamber as a result of sudden and/or abrupt closing of the culture chamber by clamping.

In accordance with a particular embodiment the fluidic device according to the invention is characterized in that the sealing means comprise a layer of adhesive material between the opposing surfaces of the upper edge of the side wall and the top wall. The adhesive material may be chosen or selected from known suitable adhesives to provide an adhesion between the top wall and side wall of the device that is sufficiently strong to maintain the top wall fixed on the side wall under normal circumstances while allowing the top wall to be manually removed from the side wall, e.g. lifting the top wall by hand.

A preferred embodiment of the fluidic device according to the invention is characterized in that the layer of adhesive material is formed by a protein precipitate, particularly a protein precipitate formed from the cell culture medium. Accordingly, the top wall and side wall can be sealed together by provision of a protein precipitate between the opposing surfaces. In a simple and preferable way this can be achieved by allowing some of the cell culture medium supplied to the chamber to be kept between the opposing edges for sufficient time to form the protein precipitate after evaporation of excess fluid. It has been found that most conventional cell culture media comprise the necessary proteins and other ‘sticky’ compounds to render a precipitate thereof sufficiently adhesive in most cases for securing the top wall to the side wall. The cell culture medium can purposively be provided between the opposing edges either directly or indirectly, for example by allowing a fluid level of the cell culture medium in the chamber to be initially higher than the upper edge of the side wall so that an amount of the cell culture medium will flow on, and possibly over, the surface of the upper edge to provide the protein precipitate after evaporation of excess fluid. A stickiness, i.e. adhesive strength, of the protein precipitate will increase over time as more of the fluid of the culture medium evaporates to a stickiness which has been found to be sufficient for a reliable sealing of the top wall on the side wall to render the chamber of the fluidic device covered and sealed from the outside environment while a maximum stickiness of the protein precipitate will allow manual removal of the top wall from te side wall when desired.

In a particular embodiment the fluidic device according to the invention is characterized in that the sealing means comprise a vacuum coupling between the opposing surfaces of the upper edge and the top wall, particularly a cavity enclosed by the opposing surfaces of the upper edge and the top wall when the upper side of the cell culture chamber is closed by the top wall, which cavity is communicatively coupled with a gas outlet configured for coupling to fluid pumping means. Such vacuum coupling allows for reversible sealing of the top wall to the side wall by pumping a gas, e.g. air, out of the cavity respectively to the cavity enclosed between the opposing surfaces of the upper edge and the top wall. Such vacuum sealing may be applied in the fluidic device as alternative sealing means or as additional sealing means to the layer of adhesive material described in the foregoing.

In accordance with a particular embodiment the fluidic device according to the invention is characterized in that the fluidic channel is a microfluidic channel. In a microfluidic channel properties of a fluid flow are very predictable as turbulence in the flow is minimized. Accordingly a flow of fluids as well as compounds and substances carried by such fluids to the cell culture can be reliably controlled under microfluidic circumstances, so as to be able to determine exact quantities being supplied to the cells. The fluidic channel preferably has a size, and particularly a cross section, mimicking the size and cross section of in vivo vascular vessels, e.g. lymphatic vessels or blood vessels such as arteries, arterioles, veins, venules and capillaries. The fluidic channel may also have other sizes and/or cross sections. The channel may for example have an cross section between 0.05 - 10 mm, wherein a channel with a cross section below 0.5 mm represents a microfluidic channel.

Preferably the opening in the bottom wall of the fluidic device through which the chamber and the fluidic channel are in fluid communication is arranged to prevent cells from migrating from one side of the opening to the opposite side of the opening while allowing a flow of fluids and compounds between the chamber and the fluidic channel. Preferably the opening is a micro-opening and has a largest cross section area of between 0.05 mm² - 1 mm².

A further preferred embodiment of the fluidic device according to the invention has the chamber comprising an upper chamber section arranged as a cell culture medium reservoir at the upper side of the chamber and a lower chamber section arranged at the bottom side of the chamber for maintaining and culturing cells, the lower chamber section being in fluid communication with the fluidic channel through the opening in the bottom wall. By providing the chamber with a reservoir on the upper side a relatively large amount of culture medium can be provided on top of the cell culture provided in the bottom section of the chamber, so that the device can be used in a closed state of the chamber for a longer duration before the amount of culture medium becomes a limiting factor in growth and/or maintenance of the cell culture. As such the chamber is preferably completely filled with culture medium prior to covering the chamber by placing the top wall on top of the upper edge of the side wall to provide the largest possible amount of fresh medium. By providing the medium, such as a fluid, in the chamber on top of the lower chamber section, a medium pressure, i.e. static fluid pressure of the medium, is applied on a cell culture that is provided in the lower chamber section, thereby stimulating perfusion of the fluid. Moreover an initially excessive amount of culture medium supplied to the chamber, i.e. a volume of the supplied medium being larger than the internal volume of the chamber, will provide a fluid level of the cell culture medium in the chamber to be higher than the upper edge of the side wall so that an amount of the cell culture medium will flow on the surface of the upper edge, which as is described in the foregoing is a convenient way of providing a protein precipitate as sealing means between the top wall and side wall of the device. Preferably a head space in the chamber above the supplied culture medium is prevented as such head space could in use of the device in the closed state provide less clear view through the top wall, e.g. due to condensation on the surface of the top wall facing the chamber, resulting in less than optimal monitoring of cells with high spatial and temporal resolution.

In a particular embodiment of the fluidic device according to the invention the lower chamber section is formed by a recess in the bottom wall. For instance during manufacturing of the fluidic device the lower chamber section can be simply provided in the bottom wall by suitable molding.

Preferably in an embodiment of the fluidic device according to the invention a plurality of recesses and openings is provided in the bottom wall with each recess forming a lower chamber section and being in fluid communication with the fluidic channel through a respective opening of the plurality of openings in the bottom wall. Separate cell cultures of the same type of cells or of different types of cells can be maintained in corresponding lower chamber section of the plurality of lower chamber sections, while the device has a single upper chamber section forming the cell culture medium reservoir which supplies each of the separate cell cultures.

The plurality of recesses may be provided as a series of lower chamber sections extending in a direction of the fluidic channel and in fluidic communication therewith through the respective opening. The series of lower chamber sections may be used to represent a network of cells or cell tissue. Particularly a gradient of one or more compounds can be supplied to the series of lower chamber sections via the fluidic channel, with a lower chamber section in fluid communication closest to the inlet of the fluidic channel being supplied with the highest concentration of compound and a lower chamber section in fluid communication closest to the outlet of the fluidic channel being supplied with the lowest concentration of compound.

In a particular embodiment of the fluidic device according to the invention at least one further fluidic channel is provided to form a set of fluidic channels in the device. A set of fluidic channels in the device allows for the supply of fluid or components of the fluid to the chamber at different locations, thus mimicking a network of supply vessels such as blood vessels and capillary lymph channels for a tissue. Different or identical fluids may be flown through each fluidic channel in order to supply components of interest such as nutrients, chemicals, signaling proteins and/or other biomolecules and factors, to the cell culture in the chamber.

In a further particular embodiment of the fluidic device according to the invention a set of fluidic channels is provided, each channel extending between a respective channel inlet and channel outlet. A further preferred embodiment of the fluidic device according to the invention has fluidic channels of the set of fluidic channels extending parallel to each other, with each fluidic channel being in fluid communication with at least one respective lower chamber section, and in particular with a respective series of lower chamber sections.

A further preferred embodiment of the fluidic device according to the invention is characterized in that between the fluidic channel and the cell culture chamber a cell transport barrier is arranged, and wherein the cell transport barrier particularly comprises a membrane. Preferably the cell transport barrier is provided in the opening or openings of the bottom wall.

The cell cultures may be provided on a suitable surface of the device according to the invention, e.g. on a surface of the bottom wall facing the chamber, for forming for example a 2D cell layer on the surface. In a particular embodiment the fluidic device according to the invention is provided with a cell scaffolding substance, particularly a gel, on either or both sides of the cell transport barrier for maintaining cells in a 3D cell culture. The cell scaffolding substance may form a fluid flow barrier to further restrict a fluid flow, possibly in addition to the cell transport barrier. The scaffolding may be used to redirect a free flow of a fluid to a certain extent. The scaffolding may be a complete restriction in that no flow of fluid there through is allowed, or may be a partial restriction in that some fluid flow there through is possible. The scaffolding may in any event allow movement of the fluid or parts thereof, for example compounds, particles, or other substances and/or components in the fluid, by means of diffusion. Because of the provision of a scaffolding substance in the chamber or fluidic channel direct fluid exchange there between is at least restricted, i.e. separating the fluid channel to some extent from the chamber. Thus, any fluid flowing through the fluidic channel in the fluidic device is for a larger part directed along the chamber, preventing a strong flow of such fluid into and/or through the chamber. As a consequence only cells in the chamber exposed on an outside of the scaffolding substance facing the fluidic channel will possibly experience shear stress across the surface, whereas this will not or hardly be the case for a cell culture captured in the scaffolding substance more remote from the fluidic channel.

The provision of the cell scaffolding substance as fluid flow barrier may be used to mimic the mechanical forces that help govern the architecture of tissues such as vascular tissues. Importantly many cell types including fibroblasts, smooth muscle cells, osteocytes, and chondrocytes, reside within a three-dimensional environment and are exposed to interstitial fluid forces. Physiological interstitial flow is the movement of fluid through the extracellular matrix of a tissue, often between blood vessels and lymphatic capillaries. It provides convection necessary for the transport of large proteins through the interstitial space and constitutes an important component of the microcirculation. Interstitial flow also provides a specific mechanical environment to cells in the interstitium that could play an important role in determining interstitial organization and architecture. Thus the fluidic device according to the invention may employ a scaffolding substance that forms a fluid flow barrier between the fluidic channel and the culture chamber which allows for a flow of fluid from the fluidic channel through the scaffolding substance mimicking that of interstitial flow, in order to expose the cultured cells to interstitial fluid forces and to provide the cells with the necessary or intended nutrients and/or other biomolecules. Types of cells, such as endothelial and epithelial cells, that in tissue form a monolayer to create a lumen or surface and are exposed to shear stresses across the surface, may be seeded on the outside of the scaffolding substance to be exposed to the fluid flow of fluid moving through the fluidic channel of the device.

A further preferred embodiment of the fluidic device according to the invention is characterized in that the upper edge of the side wall and the top wall at the opposing surfaces are made of different material and at least one of the upper edge and the top wall is arranged to elastically deform at the opposing surfaces when an external force is applied thereon. The elastic deformation improves the durability of the fluidic device and minimizes wear or other possible deterioration of the device after repeatedly opening and closing of the chamber by replacing the top wall on the side wall.

A further preferred embodiment of the fluidic device according to the invention is characterized in that the bottom wall and side wall are formed as an integral body, and particularly form a monolithic chip. The top wall may also be an integral part of the fluidic device, but is preferably formed by a separate body which can be completely detached from the bottom wall and side wall.

In another embodiment the fluidic device according to the invention is characterized in that the integral body, particularly monolithic chip, is made from PDMS. Thus the integral body, preferably formed of the bottom wall and side wall may be formed e.g. by molding.

A further preferred embodiment of the fluidic device according to the invention is characterized in that the top wall is formed by a platedike body or plate, particularly a cover slip. Preferably, the platedike body or plate, particularly cover slip, is made from glass.

Another embodiment of the fluidic device according to the invention is characterized in that the top wall is supported at an outer edge thereof on the upper edge of the side wall when the culture chamber is closed at the upper side by the top wall. Accordingly the top wall has no or at least no substantial rim extending laterally with respect to the side wall.

A particularly preferred embodiment of the fluidic device according to the invention is characterized in that the microfluidic device is configured and intended for use in an organ-on-chip. In a further aspect a cell culturing system is provided, which cell culturing system comprises a fluidic device as described herein coupled with a fluid perfusion device configured for providing a fluid flow through the fluidic channel between the channel inlet and channel outlet wherein fluid from the fluid flow can perfuse through the opening into the open-top chamber.

A preferred embodiment of the cell culturing system according to the invention has the fluid perfusion device comprising a pump arranged for pumping the fluid through the fluidic channel with predetermined physiological volumetric flow rates, particularly volumetric flow rates mimicking the in vivo volumetric flow rates of human blood stream.

In a further particular embodiment of the cell culture system according to the invention the fluidic device is provided with tissue cells in cell culture medium contained in the chamber and endothelial cells contained in the fluidic channel. Preferably the provided tissue cells are cardiac cells.

In another aspect a method of testing a compound, particularly a drug, on a cell culture, particularly a 3D cell culture, is provided, in which method a fluidic device as described herein is used and/or a cell culturing system as described herein is used, the method comprising filling of the open-top chamber with cell culture medium through the open access upper side of the chamber, seeding cells in the culture medium, placing the top wall on the upper edge of the side wall to close the chamber at the upper side, and supplying the compound to the cell culture via the fluidic channel.

In accordance with some embodiments, a method for culturing cells, tissue, or both cells and tissue, or co-culturing cells and tissue includes using a fluidic device described herein.

The "fluidic channel" used herein refers to a path of fluid flow. In some cases, a fluid path defines a space in which cells or tissue is cultured and which is open at both sides to be connected with another flow path or chamber so as to allow exchange of a culture medium and a fluid between adjacent fluids or chambers.

The fluidic channel as used herein may have the same cross section along its length between the channel inlet and channel outlet, or may instead have a varying cross section along its length between the channel inlet and channel outlet. The fluidic channel and any other channel in the device may have a cross section below 0.5 mm, thus representing microfluidic channels.

The “fluidic device” as used herein may be a microfluidic device, i.e. comprises one or more space such as a channel or chamber having small-scale or micro-scale dimensions, particularly sub -millimeter scale, in which one or more spaces a fluid flow has microfluidic properties. The fluidic device may in addition or alternatively comprise one or more macro-scale spaces in which a fluid flow displays macrofluidic behavior. The iluidic device may typically be a microfluidic chip.

The terms "recess", "chamber" or "reservoir" used herein refer to a space for containing a culture medium to culture cells or tissue.

Brief description of the drawing

For a better understanding of the various described aspects and embodiments, reference is made to the appended drawing in which like reference numerals refer to corresponding parts throughout the figures. The drawing is not in any way meant to reflect a limitation of the scope of the invention, unless this is clearly and explicitly indicated. In the drawing:

Fig. la depicts a perspective view of an embodiment of a iluidic device according to the invention.

Fig. lb depicts a detailed view of a cross section of the embodiment of the iluidic device of fig. la viewed along line A- A as illustrated in fig. la.

As shown in Fig. la and Fig. lb the iluidic device 1 has a bottom wall 2 defining a bottom side of an open-top chamber 3 arranged for containing a cell culture medium. A pair of parallel iluidic channels (not shown) extends below the chamber, each iluidic channel extending between a respective channel inlet 4a, 4b and a channel outlet 5a, 5b for guiding a fluid flow. The iluidic channels may be provided in the bottom wall, for example by suitable molding of at least one of two separate lower wall parts (not shown). A side wall 6 of the iluidic device extends from the bottom wall 2 circumferentially around the chamber 3 and has an upper edge 7 delimiting an open access upper side of the chamber 3. The bottom wall 2 and side wall 6 form a molded integral body made of PDMS. In use of the iluidic device a top wall (not shown) may be provided on and supported by the upper edge 7 for closing the chamber at the upper side thereof. The top wall is preferably a relatively simple plate-like body, such as a coverslip, having a uniform thickness. The top wall is further preferably transparent so as to allow visual inspection of the culture chamber when the top wall covers the chamber. The top wall may for instance be made of PDMS or glass. In order to shield the cell culture and the culture conditions from possible outside influences the top wall is sealed to the upper edge 7 in use of the fluidic device. This may be realized by any suitable sealing means, such as vacuum sealing via a vacuum coupling between the opposing surfaces of the upper edge and the top wall which allows for reversible sealing of the top wall to the side wall by pumping a gas, e.g. air, between the opposing surfaces of the upper edge and the top wall. Preferably the sealing means comprise a layer of adhesive material between the opposing surfaces of the upper edge and the top wall. The layer of adhesive material is preferably formed in use of the device by supplying culture medium to the culture chamber in excess, i.e. completely filling the total volume of the culture chamber with culture medium such that an excessive amount of culture medium flows over the upper edge 7 when the top wall is placed over the culture chamber. The excessive amount of culture medium flowing over the upper edge will over time by liquid evaporation form an adhesive precipitate between the surfaces of the upper edge and the top wall that seals the top wall to the side wall under normal device operative circumstances. Such sealing by adhesive forces has been found to be adequate and suitable in use of the device to shield the cell culture and the culture conditions from possible outside influences, and particularly provides a fluid tight closure of the culture chamber to the outside environment. An unexpected and/or undesired movement of the top wall with respect to the side wall is under such normal operative circumstances prevented, while the adhesion strength can be easily overcome by applying a manual lifting or sliding force on the top wall which allows the top wall to be moved away to enable access to the culture chamber when desired, e.g. to retrieve cells from a cell culture in the culture chamber.

As shown in figures la and lb the chamber 3 comprises a relatively large volume upper chamber section 10 defined by a top surface of the bottom wall 2 and an inner surface of the side wall 6 which upper chamber section forms a cell culture medium reservoir. The chamber 3 further comprises a plurality of recesses 20 in the bottom wall each recess forming a lower chamber section of the chamber 3 for maintaining and culturing cells. Each of the recesses 20 is in fluid communication with the upper chamber section 10 so that culture medium can be exchanged between said chamber sections. The chamber 3, i.e. the upper chamber section 10 and all recesses 20, is preferably completely filled with culture medium prior to covering the chamber with the top wall to provide the largest possible amount of fresh medium. By providing the medium, such as a fluid, in the chamber on top of the lower chamber sections, a medium pressure, i.e. static fluid pressure of the medium, is applied on a cell culture that is provided in the lower chamber section, thereby stimulating perfusion of fluids supplied.

The plurality of recesses 20 are provided in two series of recesses, each series of recesses being in fluid communication with one of the pair of fluidic channels via respective openings 8 provided in the bottom wall between each recess and the respective channel so that each of the recesses can be supplied with fluid via the respective fluidic channel. The opening 8 of each recess is preferably a slit in the bottom wall, which slit is arranged to prevent cells from migrating from one side of the opening to the opposite side of the opening while allowing a flow of fluids and compounds between the recess and the fluidic channel. Preferably the opening is a micro-opening and has a largest cross section area of between 0.05 mm² - 1 mm².

Separate cell cultures of the same type of cells or of different types of cells can be maintained in corresponding recesses of the plurality of recesses. Different or identical fluids may be flown through each fluidic channel in order to supply components of interest such as nutrients, chemicals, signaling proteins and/or other biomolecules and factors, to the cell cultures in each of the recesses corresponding to the fluidic channel concerned. Preferably the fluidic device is provided with cardiac tissue cells as a 3D tissue in each recess 20 with suitable cell culture medium being provided in the chamber 3, and endothelial cells being provided as a monolayer tissue in each fluidic channel to mimic a vessel and/or capillary, so that the fluidic device is particularly suited to experiment and study perfusion, diffusion and/or other exchange of compounds between the cell structures resembling in vivo conditions. For the purpose of clarity and a concise description features are described herein as part of the same or separate embodiments, however, it will be appreciated that the scope of the invention may include embodiments having combinations of all or some of the features described throughout the application. It will be clear to the skilled person that the invention is not limited to any embodiment herein described and that modifications are possible which may be considered within the scope of the appended claims. Additions, deletions, and modifications within the purview of the skilled person may generally be made without departing from the scope of the invention, as determined by the claims.

Claims

1. A fluidic device, in particular a microfluidic chip, for culturing a 3D cell culture comprising:

- a bottom wall defining a bottom side of an open-top chamber arranged for containing a cell culture medium wherein a fluidic channel extends below the chamber between a channel inlet and a channel outlet for guiding a fluid flow, wherein the bottom wall is provided with an opening through which the chamber and the microfluidic channel are in fluid communication;

- a side wall extending from the bottom wall circumferentially around the chamber to a top wall opposite the bottom wall, an upper edge of the side wall delimiting an open access upper side of the chamber; and

- the top wall arranged for closing the chamber at the upper side thereof, the top wall being supported by the upper edge of the side wall, wherein between opposing surfaces of the upper edge of the side wall and the top wall a sealing means is provided which sealing means is arranged for reversible fluid sealing of the side wall and top wall when the chamber is closed at the upper side by the top wall.

2. Fluidic device according to claim 1, wherein the sealing means comprise a layer of adhesive material between the opposing surfaces of the upper edge of the side wall and the top wall.

3. Fluidic device according to claim 2, wherein the layer of adhesive material is formed by a protein precipitate, particularly a protein precipitate formed from the cell culture medium.

4. Fluidic device according to claim 1, wherein the sealing means comprise a vacuum coupling between the opposing surfaces of the upper edge and the top wall, particularly a cavity enclosed by the opposing surfaces of the upper edge and the top wall when the upper side of the cell culture chamber is closed by the top wall, which cavity is communicatively coupled with a gas outlet configured for coupling to fluid pumping means.

5. Fluidic device according to any one of the foregoing claims, wherein the culture chamber comprises an upper chamber section arranged as a cell culture medium reservoir at the upper side of the cell culture chamber and a lower chamber section arranged at the bottom side of the cell culture chamber for maintaining cells, the lower chamber section being in fluid communication with the microfluidic channel through the opening in the bottom wall.

6. Fluidic device according to claim 5, wherein the lower chamber section is formed by a recess in the bottom wall.

7. Fluidic device according to claim 6, wherein a plurality of recesses and openings is provided in the bottom wall with each recess forming a lower chamber section and being in fluid communication with the microfluidic channel through a respective opening of the plurality of openings in the bottom wall.

8. Fluidic device according to any one of the foregoing claims, wherein between the microfluidic channel and the cell culture chamber a cell transport barrier is arranged.

9. Fluidic device according to claim 8, wherein the cell transport barrier comprises a membrane.

10. Fluidic device according to claim 8 or claim 9, wherein the cell transport barrier is provided in the opening or openings of the bottom wall.

11. Fluidic device according to any one of claims 8 - 10, wherein a cell scaffolding substance, particularly a gel, is provided on either or both sides of the cell transport barrier for maintaining cells in a 3D cell culture.

12. Fluidic device according to any one of the foregoing claims, wherein the upper edge of the side wall and the top wall at the opposing surfaces are made of different material and at least one of the upper edge and the top wall is arranged to elastically deform at the opposing surfaces when an external force is applied thereon.

13. Fluidic device according to any one of the foregoing claims, wherein the bottom wall and side wall are formed as an integral body, and particularly form a monolithic chip .

14. Fluidic device according to claim 13, wherein the integral body, particularly monolithic chip, is made from PDMS.

15. Fluidic device according to any one of the foregoing claims, wherein the top wall is formed by a platedike body or plate, particularly a cover slip.

16. Fluidic device according to claim 15, wherein the platedike body or plate, particularly cover slip, is made from glass.

17. Fluidic device according to any one of the foregoing claims, wherein the top wall is supported at an outer edge thereof on the upper edge of the side wall when the culture chamber is closed at the upper side by the top wall.

18. Fluidic device according to any one of the foregoing claims, wherein the microfluidic device is configured and intended for use as an organ-on-chip.

19. Cell culturing system comprising a fluidic device according to any one of the foregoing claims coupled with a fluid perfusion device configured for providing a fluid flow through the channel between the channel inlet and channel outlet wherein fluid from the fluid flow can perfuse through the opening of the bottom wall into the culture chamber.

20. Cell culturing system according to claim 19, wherein the fluid perfusion device comprises a pump arranged for pumping the fluid through the channel with predetermined physiological volumetric flow rates, particularly volumetric flow rates mimicking the in vivo volumetric flow rates of human blood stream.

21. Cell culture system according to claim 19 or claim 20, comprising tissue cells in cell culture medium contained in the cell culture chamber and endothelial cells contained in the microfluidic channel.

22. Cell culture system according to claim 21, wherein the tissue cells are cardiac cells.

23. Method of testing a compound, particularly a drug, on a cell culture, particularly a 3D cell culture, using a fluidic device according to any one of claims 1 - 18, and/or a cell culturing system according to any one of claims 19 - 20, comprising providing the cell culture chamber with cell culture medium through the open access upper side of the cell culture chamber, seeding cells in the culture medium, placing the top wall on the upper edge of the side wall to close the cell culture chamber at the upper side, and supplying the compound to the cell culture via the fluidic channel.