WO2011073536A1

WO2011073536A1 - Platform system, elements and use thereof, and manufacturing methods

Info

Publication number: WO2011073536A1
Application number: PCT/FI2010/051063
Authority: WO
Inventors: Marjo Yliperttula; Arto Urtti; Madhushree Bhattacharya; Jouko Peltonen; Helka Juvonen; Tapio MÄKELÄ
Original assignee: Helsingin Yliopisto; Åbo Akademi
Priority date: 2009-12-18
Filing date: 2010-12-20
Publication date: 2011-06-23
Also published as: FI20096368A0

Abstract

A platform system includes a bottom element (1) com- prising a bottom substrate plate (3) having an array of cell culture sites (5) thereon for introducing and culturing cells (9), and a cover element (10) comprising a support substrate plate (12) having an array of test agent sites (14) thereon. The cell culture sites (5) and the test agent sites (14) are bounded by walls (7, 15) formed by patterned layers(2, 11) of a hydro- phobic material printed on the substrate plates(3,2).

Description

PLATFORM SYSTEM, ELEMENTS AND USE THEREOF, AND MANUFACTURING METHODS

FIELD OF THE INVENTION

The present invention relates to means and methods for different kinds of cell culture based studies of e.g. drugs and other chemicals. Particularly, the present invention relates to the so called high throughput screening methods where a great number of test com^¬ pounds and their interactions with cells can be inves^¬ tigated at a same time. The present invention is fo^¬ cused on a platform system and elements thereof suit^¬ able for such screening and manufacturing and use of such platform system. The present invention is also focused on manufacturing and use of such platform system.

BACKGROUND OF THE INVENTION

Cell culture based screening methods for investigating the influences of different compounds on cells are used e.g. in toxicity screening of chemical compounds and drug research. The basic principle is to arrange a two dimensional array of localized cell cultures on a substrate, to bring the compounds to be investigated in contact with the cell cultures, and to observe the results of the compound-cell interactions usually visually, preferably by means of an automatic observa^¬ tion equipment capable of rapid and reliable evalua^¬ tion. One essential point in such screening assays is said arranging of the cell culture array on a sub^¬ strate .

A common solution for the substrate for a cell culture array is a so called well plate. A typical well plate consists of a transparent substrate plate with a two dimensional array of identical wells formed therein. The prior art well plates are usually manufactured by injection molding. An array of cell cultures is achieved by introducing cells together with suitable biomaterial to the wells. The compounds to be investi^¬ gated are then inserted into the wells. The cells as well as the compounds are introduced to the wells by pipetting .

Despite their many advantageous properties, the prior art well plates also suffer from many deficiencies. For example, well plates are commercially available with a limited range of sizes only. They are also re^¬ latively expensive. Possibilities to treat the well surfaces for optimizing e.g. their adhesive properties or the growth conditions of cells introduced in the wells are rather limited. The volume of a single well in a typical well plate is rather large which means that both the cells and their growth support materials and the actual drugs or other compounds to be investi^¬ gated are consumed in rather large amounts in every assay. On the other hand, introducing the cells and the growth support biomaterial as well as the test agents to the wells by pipetting is time-consuming and laborious .

Another standard solution is to form the cell array by introducing the cells and the growth supporting biomaterial directly on a glass plate substrate by locating them on the nodes of the desired array geometry. This necessitates the use of a so called plotter capable of sufficiently accurate positioning. Plotters of this kind are very expensive apparatuses available only in well equipped special laboratories. Moreover, in this kind of cell array on a substrate plate, there are no mechanical borders between the different cell cul^¬ tures. This means that the drug or other compound in^¬ troduced into a cell culture at one node of the array can be in contact also with the cells of the neighbor- ing cell cultures. Thus, the concentration of the test agent in the cell culture is difficult to control. Different test agents introduced to different cell cultures can also mix with each other in an undesirable way.

PURPOSE OF THE INVENTION

The purpose of the present invention is to disclose a platform system and elements for such platform system as well as a manufacturing method and use thereof based on an entirely new approach and providing great advantages in cell array based assays.

SUMMARY OF THE INVENTION

The present invention is characterized by what is pre^¬ sented in claims 1, 5, 6, 7, 11, and 124.

One key principle in the present invention is utiliza^¬ tion of a printing technique. Printing is to be understood here in a broad sense covering all methods where a three dimensional structure according to a predeter^¬ mined pattern is formed on a surface of an existing substrate, formed of a substantially solid material, using a printing member. Three-dimensionality means here that the structure extends not only in the plane of the substrate surface but has also a dimension in a direction perpendicular to it. However, the thickness of the structure in this direction can be very small. For example, when printing cells, minimal thickness of the printed structure is as low as the height of one layer of cells on a substrate.

In one approach of the printing as determined here, forming the three dimensional structure is performed by introducing additional material on the substrate in a predetermined pattern. In this approach, the print- ing member brings the material to the substrate sur^¬ face directly to the locations according to said pre^¬ determined pattern geometry. Typically, this is per^¬ formed by bringing the printing member in mechanical contact or at least in very close proximity with the substrate, after which the material to be printed is transferred from the printing member to the substrate. Sometimes a direct contact is not needed. This is the case e.g. in ink jet printing where the material to be printed is transferred from a printing head to the substrate as a jet of said material.

On the other hand, printing is intended to cover here also methods wherein the three-dimensional modifica^¬ tion of the substrate surface is achieved by directly pressing the three dimensional structure on the sub^¬ strate surface, i.e. without bringing any material from outside but by forming the structure in the sub^¬ strate material itself.

Common for both approaches above - forming the desired geometry with added material and pressing it directly in the substrate itself - is that the structure is formed on an existing substrate. Compared to e.g. the prior art molding techniques, this opens a great vari^¬ ety of novel possibilities in cell culture platform system manufacturing, as is discussed in more detail later in this document.

Known printing techniques suitable for the present in^¬ vention include e.g. inkjet printing, rotogravure, flexography, reverse-gravure coating, hot embossing, offset printing, and screen printing. However, no printing technique is excluded.

It is possible that for the purposes of the present invention, the printing processes usually used for producing other types of products, e.g. graphical products, have to be modified somehow. However, by proper selection of the actual printing technique, compatibility between the process and the printing ma^¬ terial according to the present invention is often achievable by adjusting the material properties, e.g. its rheological and solution chemical properties.

Common advantages for the printing techniques meant here include cost-efficiency already in rather small production volumes, versatility of the manufacturing process enabling very flexible production, and accu^¬ racy of the structure produced.

According to one aspect, the present invention is fo^¬ cused on a bottom element to be used e.g. in cell cul^¬ ture screening. Cell culture screening relates to methods for studying effects of different test agents, e.g. drugs or other chemicals or genetic modifiers, on living cells. In typical cell culture screening, a large amount of cell cultures are arranged as an ar^¬ ray. Test agents are introduced in contact with the cell cultures and the results of the interaction be^¬ tween the test agents and the cells are then observed, usually optically by automatic observing devices. The expression "screening" refers to the possibility to rapidly and efficiently scan through the cell culture array, usually abbreviated simply as "cell array", in order to screen whether or not a particular effect is observed in each of the cell cultures. By screening methods of this kind it is possible to investigate the effects of different test agents to one particular type of cells as well as the effect of a particular test agent on different cell types.

The bottom element of the present invention comprises a bottom substrate plate having an array of cell cul- ture sites, i.e. locations for cell cultures, thereon for introducing and culturing cells.

According to the present invention, the cell culture sites are bounded by walls formed by a first patterned layer of a hydrophobic material printed on the bottom substrate plate. This principle opens a great variety of entirely new possibilities. The hydrophobic walls around the cell culture site effectively define the area on the bottom substrate plate surface on which the cells can adhere. This is a great advantage in comparison to the prior art well plates where the lo^¬ cations of the cell adherence and cell culture forma^¬ tion are defined mechanically only by the walls of the wells. Moreover, a printed layer enables a very accu^¬ rate geometry of the structure also with sizes of the cell culture sites well below those of the prior art well plates. In many but not all applications it is preferable that the patterned layer of a hydrophobic material is continuous covering the entire bottom sub^¬ strate plate except for the cell culture sites. This minimizes the possibilities for adherence of cells as well as the possible biomaterial described below else^¬ where than to the cell culture sites.

In one preferred embodiment, a biomaterial bedding is placed at a cell culture site on the bottom substrate plate to promote cell culture formation therein. Bio^¬ material means here any ingredient enhancing the ad^¬ herence and/or growth of cells on the cell culture sites. The biomaterial can be any synthetic or nature originating polymer having these kinds of properties. It can also be some extracellular living organism material of a biological origin, examples of these in^¬ cluding proteoglycans, different growth factors, and collagen. In addition to the biomaterial, also siRNA molecules, possibly of different types in different cell culture sites, can be arranged in the cell cul^¬ ture sites to knock down the target gene expression. Preferably, the biomaterial bedding is formed by printing. This ensures again very accurately located material ensuring finally high similarity between the cell cultures in different cell culture sites.

Said advantage of a possibility to have small-area and small-volume cell culture sites is particularly clear in an embodiment of the present invention where the cell culture site width lies in the range of 0.1 - 1.0 mm and the height of the wall bounding the cell cul^¬ ture site is equal to or less than 100 μιη. This kind of a bottom element can be called a "micro well plate". By the width and the height is meant here the average values of the cell culture sites of a single bottom element, which are usually substantially iden^¬ tical with possibly slight variation. The width means here the characteristic width of a cell culture site, e.g. the diameter in the case of a circular shape and the length of the longer side in the case of a rectan^¬ gular one. This small size of the cell culture sites dramatically reduces the necessitated size of the cell cultures and particularly the amounts of the biomate^¬ rial as well as the actual test agent needed. Thus, this embodiment combines efficiently the best proper^¬ ties of both of the most common prior art solutions, i.e. the mechanically restricted borders of the cell culture sites present in the prior art well plates, and the very small amounts of the cells and other sub^¬ stances needed in the cell arrays plotted directly on a glass substrate. It also enables three dimensional cell cultures with a reasonable number of cells. Three- dimensionality is important in the sense that most of the tissues of interest in typical screening assays are three dimensional. Thus, results from an assay performed with two dimensional cell cultures do not truly correspond to the situation within a living body and can thus lead in false conclusions e.g. in drug development.

On the other hand, in a bottom element according to the present invention, the well size can also be sub^¬ stantially larger if needed; the printing enables very flexible adjusting of the patterned layer structure.

In a preferred embodiment, the bottom element com^¬ prises fluid channels opening into the volumes at the cell culture sites, the volumes being defined by the bottom substrate plate surface and the walls bounding the cell culture sites, for supplying fluid to the cell cultures formed therein. Fluid channels can be used e.g. for adding sustaining solution for the cells in order to keep them alive.

To form a cell array using a bottom element according to the present invention, cells are introduced into the cell culture sites within a suitable matrix pre^¬ serving the cells living. This can be performed by means of an automatic pipetting or plotting equipment. Alternatively, the cells can be brought all over the bottom element and then rinsed, whereupon excess cells outside the cell culture sites are removed. On the other hand, it is possible to use the basic principle of utilizing a printing technique, e.g. flexography, also in introducing the cells to the cell culture sites .

In order to enable a complete platform system, another aspect of the present invention is focused on a cover element stackable on a bottom element as defined above "upside down" for introducing one or more test agents in contact with cell cultures formed in the cell cul^¬ ture sites of the bottom element. The cover element comprises a support substrate plate having a test agent site array thereon, the test agent sites being bounded by walls formed by a second patterned layer of a hydrophobic material printed on the support sub^¬ strate plate. One or more immobilized, releasable test agents are inserted in the volumes at the test agent sites, the volumes being defined by the support sub^¬ strate plate surface and the walls bounding the test agent sites. A removable protective membrane lies on the second patterned layer of a hydrophobic material, the protective membrane closing said volumes defined by the support substrate plate surface and the walls bounding them.

The cover element has thus a structure with a basic construction similar to that of the bottom element, i.e a substrate plate wherein an array of hollows de^¬ fined by the substrate surface and a patterned layer of a hydrophobic material on the substrate plate is formed. In the cover element, the purpose of the hol^¬ lows is to insert test agents therein. First, immobi^¬ lized means that a test agent at a test agent site is attached to some "carrier" substance, e.g. some solid polymer matrix, for keeping the test agent immovable. Releasable, in turn, means that the test agent can be controllably released from this carrier when the cover member is stacked on a bottom member. The release can be controlled e.g. by erosion or osmotic imbibitions of water into the test agent site. It is also possible to use an osmotic pump integrated in the platform sys^¬ tem. As one possibility, soft battery technology can be used to activate the test agent release. In this case, soft batteries are integrated in the bottom and cover elements and iontophoresis is used to transfer the drugs to the cells in order to increase the recov^¬ ery of drugs into the cells. Moreover, the geometry of the array of test agent sites of the cover element corresponds to the geometry of the array of cell culture sites of the bottom ele^¬ ment for providing coincidence between the cell cul^¬ ture sites and the test agent sites when stacking the cover element on the bottom element with the side of the protective member against the bottom element, i.e. "the upside down". By coincidence between the test agent sites and the cell culture sites is meant here that when stacking the bottom element and the cover element that way, those "wells" of the bottom and cov^¬ er elements formed by the patterned hydrophobic mate^¬ rial layers open face to face. Further, such corre^¬ sponding or mirrored geometries mean also that the test agent site width lies preferably in a range sub^¬ stantially similar to that of the cell culture sites.

As is clear from the definition above, the purpose of the cover element is to provide a way to introduce one or more test agents in contact with the cells of a cell array formed on the bottom element efficiently and substantially simultaneously to all cell cultures. In practice, this is achieved by removing the protec^¬ tive membrane of the cover element, setting the cover element upside down on the bottom element where a cell culture array is formed in the cell culture sites, and activating the test agent release, after which the test agent is diffused to the cells.

All test agent sites of a cover member can be occupied by one particular test agent. This approach is applied when it is desired to study the effect of said test agent on different cell types. On the other hand, per^¬ haps more typically, the final purpose of a screening assay is to investigate the effects of several test agents on one or more cell types. Then, a cover member forms a kind of test agent "library". This can com- prise some standard group of agents or it can be tai^¬ lored according to specific needs of a particular study .

At least one of the bottom substrate plate and the support substrate plate, preferably both of them, is made of a transparent material enabling optical in^¬ spection of the cell-test agent interaction through the substrate plate (s).

Possible materials for the substrate plates are e.g. polymethyl methacrylate PMMA and hyarylonan. Materials suitable for the patterned layers of a hydrophobic ma^¬ terial include e.g. PMMA, hyaluronan, and silicon.

As one aspect, the inventive principle of the present invention also covers an entire platform system comprising both a bottom element and a cover element as defined above.

On the other hand, the present invention is also fo^¬ cused on a method for manufacturing a bottom element comprising a bottom substrate plate having an array of cell culture sites thereon for introducing and cultur- ing cells. The principles of this kind of a bottom element are described above.

According to the present invention, the method comprises printing a first patterned layer of a hydropho^¬ bic material on the bottom substrate, the patterned layer forming walls bounding the cell culture sites.

In contrast to the prior art methods typically based on injection molding or similar techniques, in the present invention the "wells", i.e. the cell culture sites bounded by walls formed by the patterned layer of a hydrophobic material are formed by printing. This provides great advantages. First, different printing techniques enable a very accurate geometry of the pat^¬ terned layer which means that well sizes well below those of the prior art are achievable. Printing en^¬ ables also very cost efficient production already in small production volumes making it possible to flexi^¬ bly adjust the production. For example, the size of the cell culture sites can be adjusted easily. This is not the case e.g. in injection molding which is known for high costs of preparing a new mold. In addition, one very advantageous feature enabled by printing is that a large amount of bottom elements can be formed simultaneously as a large, continuous bottom element preform, from which the final bottom elements can be cut to the desired sizes. The size of the preform is practically limited by the printing equipment proper^¬ ties only.

In one preferred embodiment of the present invention, a biomaterial bedding is inserted, preferably by printing, at a cell culture site on the bottom sub^¬ strate plate to promote cell culture formation there^¬ in. In addition to the accuracy and cost efficiency, printing in its different variations enables very flexible selection of the actual composition of the biomaterial. Thus, printing offers a possibility for great variety of functionalized features to be in^¬ cluded in the cell culture sites.

Preferably, the patterned layer of a hydrophobic mate^¬ rial is printed so as to form the cell culture site width in the range of 0.1 - 1.0 mm and the height of the wall bounding the cell culture site equal to or less than 100 μιη. This small size provides advantages described above in connection with the platform system aspect of the present invention. To enhance the functionality of the platform system, in one preferred embodiment of the present invention the method comprises forming in the bottom element fluid channels opening into the volumes at the cell culture sites, the volumes being defined by the bottom substrate plate surface and the walls bounding the cell culture sites, for supplying fluid to the cell cultures formed therein. One simple way to form the fluid channels is to include the channels in the de^¬ signed pattern of the first patterned layer of a hy^¬ drophobic material which is then printed on the bottom substrate plate. The fluid channels can be open through the entire thickness of the patterned layer. Another alternative is to form the first patterned layer as two sublayers formed by introducing additive material on the bottom substrate plate, the first of which including the channels and the other one forming a cover on them.

On the other hand, one aspect of the present invention is focused on method for manufacturing a cover element stackable "upside down" on a bottom element as defined above in order to introduce one or more test agents in contact with cell cultures formed in the cell culture sites of the bottom element.

This method comprises first providing a support sub^¬ strate plate having an array of test agent sites thereon. Secondly, a second patterned layer of a hy^¬ drophobic material is printed on the support substrate plate, the second patterned layer forming walls bound^¬ ing the test agent sites. Immobilized, releasable test agent is then inserted into each of the volumes at the test agent sites, the volumes being defined by the support substrate plate surface and the walls bounding the test agent sites. One single test agent can be in^¬ serted into the all volumes. Alternatively, different test agents can be inserted into different volumes, e.g. such that each volume has a unique test agent differing from the test agents of all other volumes. Finally, a removable protective membrane is formed on the layer of the hydrophobic material, the protective membrane closing said volumes.

In the method for manufacturing a cover element, the geometry of the array of test agent sites is formed to correspond to the geometry of the array of cell cul^¬ ture sites of the bottom element for providing coinci^¬ dence between the cell culture sites and the test agent sites when stacking the cover element on the bottom element with the side of the protective member against the bottom element.

The first steps of the method for manufacturing the cover element until printing the second patterned layer of a hydrophobic material can be performed ac^¬ cording to similar principles and processes as the corresponding steps in the manufacturing of the bottom element. As described earlier in this description, the test agent to be inserted to the test agent site is attached to e.g. some solid polymer matrix or other "carrier" substance. The insertion can be made e.g. by spotting. However, more preferably, in inserting the one or more immobilized, releasable test agents into the volumes at the test agent sites, at least one test agent is inserted by printing. Printing provides an alternative for special and expensive spotting equip^¬ ment. In general, printing offers also in this step the advantages of cost efficiency, accuracy, and flexibility described above.

Advantageously, in any of the embodiments of the meth^¬ ods for forming the bottom element and the cover ele^¬ ment, at least one of the method steps performed by printing is carried out as a roll-to-roll process. By roll-to-roll process it is meant a process where a substrate to be modified is led to the modification device (s) performing the actual printing as a continu^¬ ous material strip being uncoiled from a roll. After the modification device (s), the strip is again coiled on another roll. Thus, the substrate is in a continu^¬ ous movement. This general principle is well known e.g. from printing of books and newspapers. In the methods of the present invention, a roll-to-roll proc^¬ ess can be used in any of the steps performed by printing, including printing the first and the second patterned layers of the bottom element and the cover element, respectively, but also, for example, printing the biomaterial bedding. A roll-to-roll process makes it possible to produce and modify efficiently platform system preforms having lengths in a range of even tens of meters. This is an enormous step of development in comparison with e.g. the different variations of the prior art molding techniques.

Concerning the first and second patterned layers of hydrophobic material according to the present inven^¬ tion, it is important to note that they can be also formed of the same material as the bottom and the sup^¬ port substrate plates, respectively. Particularly, this is the case always when the first and the second patterned layers are formed by pressing the three di^¬ mensional structures thereof directly on the bottom or the support substrate plate material. In this case, the "patterned layer" means that portion of the sub^¬ strate plate thickness into which the three dimen^¬ sional patterns extend from the substrate plate sur^¬ face .

In the manufacturing methods according to the present invention, different printing techniques can be util- ized in different stages of the process producing a platform system. For example, ink jet could be in some applications a good choice for forming the patterned layers. By ink jet, the patterned layers could be formed, for example, as circular structures around the cell culture or test agent sites only, i.e. without any continuous layer covering the entire substrate plate at issue. Similarly, some other technique and apparatus could be used for inserting the biomaterial beddings to the cell culture sites, and so on. From a manufacturing equipment point of view, a good approach could be to arrange the different printing devices of the different manufacturing steps in an integrated hy^¬ brid equipment where the process would be performed as a continuous roll-to-roll process.

According to yet another aspect, the present invention is implemented as use of the platform system described above in a cell culture based screening assay. The cell culture based assay means here any assay where the actual analysis is focused on a plurality of cell cultures arranged as an array on a substrate. Examples of assays of this kind comprise different biologic studies, toxicological chemical screening, drug screening and burns curing. In screening, a bottom element described above is used as a platform for form^¬ ing an array of cell cultures, and a cover element as a platform for forming a test agent array for intro^¬ ducing tests agents in contact with the cell cultures. DETAILED DESCRIPTION OF THE INVENTION

The present invention is now described in more detail with reference to the accompanying figures illustrat^¬ ing one exemplary platform system and elements thereof according to the present invention as well as manufac^¬ turing and use of such a platform system.

Figure 1 illustrates the steps of manufacturing a bot^¬ tom element of the platform system and forming a cell culture array thereon.

Figure 2 illustrates the steps of manufacturing a cov^¬ er element of the platform system.

Figure 3 illustrates one phase in using the platform system.

Figure 4 shows an overall view of the platform system.

Drawings in the figures are not in scale.

The manufacturing method of a bottom element 1 illus^¬ trated in figure 1 starts by printing a first pat^¬ terned layer 2 of a hydrophobic material, e.g. PMMA, on a bottom substrate 3 formed of a transparent poly^¬ mer. The first patterned layer comprises holes 4 at the locations of predetermined sites 5 of the sub^¬ strate surface. These sites form a two dimensional ar^¬ ray. The hole diameter can be e.g. 0.5 mm. As a result of this step, the bottom element comprises an array of small open "wells", one of which being shown in the drawings of the figure.

In the example of figure 1, the patterned layer is printed as two sub-layers, the first of which compris^¬ ing also channels 6 opening to the wells through the walls 7 of the hole in the patterned layer. These channels can be used to introduce fluid into this vol^¬ ume .

As the next step, a bedding 8 of a biomaterial promot^¬ ing cell adhesion and growth is printed on the sub^¬ strate surface at said predetermined sites. Examples of possible materials for this bedding are given in the summary section of this specification.

The bottom element formed in this way can be used to form a cell culture array by introducing cells 9 to the wells of the bottom element. This can be performed e.g. by spreading cells all over the bottom element surface and then removing the excess substance by rinsing. The hydrophobic material of the patterned layer prevents cell adherence elsewhere than on the biomaterial bedding 8. On the other hand, in order to utilize the full potential of the principle of the present invention based on printing, the cells can be printed directly on the cell culture sites. As is clear for a person skilled in the art of cell cultur- ing, cells are in both cases introduced to the cell culture sites within a suitable sustaining solution.

The manufacturing process of the cover element 10, il^¬ lustrated in figure 2, starts by printing a second patterned layer 11 of a hydrophobic material on a transparent support substrate plate 12. The materials of the substrate and the patterned layer as well as the printing process details can be similar to those used in manufacturing the bottom element 1 of figure 1. The second patterned layer comprises holes 13 at the locations of predetermined sites 14 of the support substrate plate. These sites form a two dimensional array, the geometry of which is a mirror of the geome^¬ try of the corresponding array at the bottom substrate plate 3. The walls 15 of the holes in the second pat- terned layer form together with the support substrate plate surface a two dimensional array of open wells in the cover element.

Next, test agent 16 is inserted in each of the wells attached in a solid polymer matrix making the test agent substantially immobile. The insertion is pref^¬ erably made by printing. Printing enables the use of different test agents, e.g. variations of a drug com^¬ pound or some other group of different chemicals, for different wells. Finally, a protective membrane 17 is formed on top of the structure, the membrane closing the volumes at the test agent sites and protecting the test agent (s) from influences of ambient conditions and interaction with external substances.

Figure 3 illustrates how the bottom and the cover ele^¬ ments 1, 10 can be used for introducing one or more test agents in contact with the cell cultures formed on the bottom element. This is achieved by removing the protective membrane from the cover member and set^¬ ting the bottom and the cover elements one on the oth^¬ er, the free surfaces of the patterned layers against each other. Then, the test agent 16 of each of the test agent sites 14 is released, controlled e.g. by erosion, and it is diffused to the cells 9 of the cor^¬ responding cell culture site 5 of the bottom element, as illustrated by arrows marked in the figure.

Figure 4 shows an overall view of a bottom element and a cover element of the platform system. They both comprise a plate-like body where a two dimensional array of volumes for cell cultures or volumes filled with test agent (s), respectively, is arranged. The volumes of the bottom element are formed as open wells coated by biomaterial beddings. The cover element is covered by a protective membrane closing the test agent vol^¬ umes .

It is important to note that the scope of the present invention is not limited to the simplified examples of figures 1 to 4, but the actual details of devices ac^¬ cording to the present invention can freely vary within the claims.

Claims

1. A bottom element (1) comprising a bottom substrate plate (3) having an array of cell culture sites (5) thereon for introducing and culturing cells (9), characteri zed in that the cell culture sites (5) are bounded by walls (7) formed by a first pat^¬ terned layer (2) of a hydrophobic material printed on the bottom substrate plate (3) .

2. A bottom element (1) according to claim 1, characteri zed in that a biomaterial bedding (8) , preferably formed by printing, is placed at a cell culture site (5) on the bottom substrate plate to pro^¬ mote cell culture formation therein.

3. A bottom element (1) according to claim 1 or 2, characteri zed in that the cell culture site (5) width lies in the range of 0.1 - 1.0 mm and the height of the walls (7) bounding the cell culture sites is equal to or less than 100 μιη.

4. A bottom element (1) according to any of claims 1 to 3, characteri zed in that the bottom element (1) comprises fluid channels (6) opening into the vol^¬ umes at the cell culture sites, the volumes being de^¬ fined by the bottom substrate plate surface and the walls (7) bounding the cell culture sites, for supply^¬ ing fluid to the cell cultures formed therein.

5. A cover element (10) stackable on a bottom element (1) according to any of claims 1 to 4 in order to introduce one or more test agents (16) in contact with cell cultures formed at the cell culture sites of the bottom element, characteri zed in that the cover element comprises

- a support substrate plate (12) having an array of test agent sites (14) thereon, the test agent sites being bounded by walls (15) formed by a second patterned layer (11) of a hydrophobic material printed on the support substrate plate;

one or more immobilized, releasable test agents (16) inserted in the volumes at the test agent sites, the volumes being defined by the support substrate plate surface and the walls (15) bounding the test agent sites (14) ;

- a removable protective membrane (17) on the second patterned layer (11) of a hydrophobic material, the protective membrane closing said volumes defined by the support substrate plate surface and the walls (15); wherein

the geometry of the array of test agent sites (14) corresponds to the geometry of the array of cell culture sites (5) of the bottom element (1) for providing coincidence between the cell culture sites (5) and the test agent sites (14) when stacking the cover element (10) on the bottom element (1) with the side of the protective member (17) against the bottom element (1) .

6. A platform system comprising a bottom element (1) according to any of claims 1 to 4 and a cover element (10) according to claim 5.

7. A method for manufacturing a bottom element (1) comprising a bottom substrate plate (3) having an array of cell culture sites (5) thereon for introducing and culturing cells, characteri zed in that the method comprises printing a first patterned layer (2) of a hydrophobic material on the bottom substrate plate (3), the patterned layer forming walls (7) bounding the cell culture sites (5) .

8. A method according to claim 7, characteri zed in that a biomaterial bedding (8) is inserted, pref^¬ erably by printing, at the cell culture site (5) on the bottom substrate plate (3) to promote cell culture formation therein.

9. A method according to claim 7 or 8, characteri zed in that the first patterned layer (2) of a hy^¬ drophobic material is printed so as to form the cell culture site (5) width in the range of 0.1 - 1.0 mm and the height of the walls (7) bounding the cell cul^¬ ture sites equal to or less than 100 μιη.

10. A method according to any of claims 7 to 9, characteri zed in that the method comprises forming in the bottom element (1) fluid channels (6) opening into the volumes at the cell culture sites, the volumes being defined by the bottom substrate plate surface and the walls (7) bounding the cell cul^¬ ture sites, for supplying fluid to the cell cultures formed therein.

11. A method for manufacturing a cover element (10) stackable on a bottom element (1) according to any of claims 1 to 4 in order to introduce one or more test agents (16) in contact with cell cultures formed at the cell culture sites of the bottom element, the method comprising:

- providing a support substrate plate (12) having an array of test agent sites (14) thereon;

- printing a second patterned layer (11) of a hydrophobic material on the support substrate plate, the second patterned layer forming walls (15) bounding the test agent sites (14) ; - inserting one or more immobilized, releas- able test agents (16) into the volumes at the test agent sites, the volumes being defined by the support substrate plate surface and the walls (15) bounding the test agent sites deforming a removable protective membrane

(17) on the second patterned layer (11) of the hydrophobic material, the protective mem^¬ brane closing said volumes defined by the support substrate plate surface and the walls

(15) ; wherein

the geometry of the array of test agent sites (14) is formed to correspond to the ge^¬ ometry of the array of cell culture sites (5) of the bottom element (1) for providing coincidence between the cell culture sites (5) and the test agent sites (14) when stacking the cover element (10) on the bottom element

(1) with the side of the protective member against the bottom element (1) .

12. A method according to claim 11, characteri zed in that in inserting the one or more immobi^¬ lized, releasable test agents (16) into the volumes at the test agent sites, at least one of the test agents (16) is inserted by printing.

13. A method according to any of claims 7 to 12, characteri zed in that at least one of the meth^¬ od steps performed by printing is carried out as a roll-to-roll process.

14. Use of a platform system (1, 10) according to claim 6 in a screening assay for investigating interaction between cell cultures and one or more test agents .