WO2022038277A1

WO2022038277A1 - Microfabricated device, system and method comprising such device

Info

Publication number: WO2022038277A1
Application number: PCT/EP2021/073176
Authority: WO
Inventors: Hans KLEINE-BRÜGGENEY; Robert WEINGARTEN; Sebastian BÜHREN
Original assignee: Evorion Biotechnologies Gmbh
Priority date: 2020-08-20
Filing date: 2021-08-20
Publication date: 2022-02-24
Also published as: EP3957397A1; EP4200075A1

Abstract

The invention pertains to a device comprising an inlet channel, outlet channel, first channel having a hydrodynamic resistance R1, second channel having a hydrodynamic resistance R2, third channel having a hydrodynamic resistance R3, fourth channel having a hydrodynamic resistance R4, intermediate channel having a hydrodynamic resistance R0 and a positioner for positioning an object within the intermediate channel, the positioner preferably comprising a narrowing of the cross section of the intermediate channel; the device further comprising an inlet node, outlet node, first intermediate node and second intermediate node; wherein the inlet channel, second channel and fourth channel are interconnected via the inlet node, the outlet channel, first channel and third channel are interconnected via the outlet node, the first channel, second channel and intermediate channel are interconnected via the first intermediate node and the third channel, fourth channel and intermediate channel are interconnected via the second intermediate node; the device further comprising a first valve arranged at the second channel and configured to be actuated in order to control the flow rate through the second channel. Furthermore, the device comprises a second valve arranged at the fourth channel and configured to be actuated in order to control the flow rate through the fourth channel. The invention further pertains to a system and a method comprising the device according to the invention.

Description

Microfabricated device, system and method comprising such device

The invention refers to a device comprising an inlet channel, outlet channel, first channel having a hydrodynamic resistance R1 , second channel having a hydrodynamic resistance R2, third channel having a hydrodynamic resistance R3, fourth channel having a hydrodynamic resistance R4, intermediate channel having a hydrodynamic resistance RO and a positioner for positioning an object within the intermediate channel, the positioner preferably comprising a narrowing of the cross section of the intermediate channel; the device further comprising an inlet node, outlet node, first intermediate node and second intermediate node; wherein the inlet channel, second channel and fourth channel are interconnected via the inlet node, the outlet channel, first channel and third channel are interconnected via the outlet node, the first channel, second channel and intermediate channel are interconnected via the first intermediate node and the third channel, fourth channel and intermediate channel are interconnected via the second intermediate node; the device further comprising a first valve arranged at the second channel and configured to be actuated in order to control the flow rate through the second channel. Furthermore, the invention refers to a system comprising at least two such devices and a method for operating such system.

Such a device (and system) is known from WO 2019 048 713 A1. In particular, it has the purpose to treat and transport an object like a cell or particle through the device by means of a fluid. In order to control the fluid flow and fluid direction through the intermediate channel, the device according to WO 2019 048 713 A1 uses a second valve arranged at the third channel to control the flow rate through the third channel and furthermore the channels have different resistances. The first channel has a higher resistance than the third channel and the fourth channel has a higher resistance than the second channel. To achieve different resistances (i.e. value of resistance) it is in principle possible to change the cross-section of an appropriate channel. However, usually the size of cross-section is limited by the size of objects intended for transport through the channels. In terms of WO 2019 048 713 A1, an object that is removed from the intermediate channel has to pass the first channel to enter the outlet node. Thus, the resistance of the first channel can only be varied by adjusting its length as the size of the cross-section is given by the object size. Therefore, for several applications it is only possible to vary the resistance of a channel by varying its length. A longer channel causes more resistance (i.e. loss of fluid pressure) than a shorter channel. Therefore, it is necessary that the first channel is longer than the third channel and the fourth channel is longer than the second channel. This, however, requires a large footprint. This is disadvantageous since the environments in which such devices are usually used are limited by space or standards like a 96-well-plate for example. Therefore, the prior art device allows a relatively small amount of devices being integrated into such biotechnological environments.

The resistance of a channel can also depend on the shape of the cross-section of the channel. However, this shape is usually predetermined by the method of manufacturing the microfabricated devices. In particular, the cross-section is limited to a rectangular shape when the device is build layer by layer using conventional 2D lithography procedures.

Thus, the technical problem is to provide a device which allows controlling the fluid flow and fluid direction through the intermediate channel and additionally requires less space.

The problem is solved by a device according to claim 1 , the system according to claim 9, and a method according to claim 13. Advantageous embodiments are disclosed by the dependent claims, the description and the figures.

The basic idea of the invention is to provide a second valve which is arranged at the fourth channel and configured to be actuated in order to control the flow rate through the fourth channel. By shifting the second valve from the third to the fourth channel, it is possible to decrease the size of the channels. For example, the channels might have the same (small) size (or at least the difference between the sizes is smaller than that of the prior art). In particular, this advantage has an effect on the system according to the invention since it comprises at least two and preferably several such devices. This leads to a system which can comprise more devices on the same footprint compared to the known system. This means that the device density is higher. Furthermore, the higher the number of devices is, the higher the space-saving is. This significantly increases the number of objects that can be analyzed with such a system.

The device is configured in such a way that by actuating the first and/or second valve, the flow rate and flow direction of a fluid flowing through the intermediate channel can be controlled. The term “to actuate a valve” means that the valve is operated. The actuation of the valve includes the opening and closing of the valve. Preferably, actuation means closing the valve. The device is microfabricated. Preferably, the device comprises or consists of several layers. Preferably, the device is manufactured by replica molding from at least one master mold that has been fabricated using 2D, 2.5D or 3D lithography procedures such as UV lithography or multiphoton lithography. In particular the dimensions of the device and its components like the channels and valves have a size of up to 5000, 1000, 500, 100, 80, 50, 10, 1, 0,01 pm.

According to the invention, the first (second) valve is arranged at the second (fourth) channel and configured to be actuated in order to control the flow rate through the second (fourth) channel. Preferably, the first (second) valve is directly arranged at the second (fourth) channel. Particularly preferred, the first (second) valve is in direct and physical contact with the second (forth) channel. In particular, the flow rate is controlled by adjusting the cross-section of the channel, preferably the cross-section at the location of the valve. The term “flow rate” also comprises the case in which there is no flow through the channel. In this case the valve is completely closed and the flow rate is zero.

The device is configured in such a way that by providing a pressure difference between inlet channel and outlet channel, a fluid being within the device can flow. To allow a fluid flow from the inlet channel to the outlet channel a pressure at the inlet channel is higher than the pressure at the outlet channel. A fluid might be a gas or a liquid.

The term “closed” means that the valve is at least partially closed, in particular completely closed. The term “opened” means that the valve is at least partially opened, in particular completely opened. The term “partially opened” means that the valve is more opened than closed. For example, in case of a partially opened valve, the crosssection size of the channel is closer to the cross-section size of the channel when the valve is completely opened than when the valve is completely closed. The term “partially closed” means that the valve is more closed than opened. For example, in case of a partially closed valve, the cross-section size of the channel is closer to the cross-section size of the channel when the valve is completely closed than when the valve is completely opened.

The term that a valve is arranged at an appropriate channel could for example mean that the valve is arranged outside the channel. In particular, the valve could generate a pressure on the channel wall thereby decreasing its cross-section. Such valve and its functioning is described in WO 2019 048 713 A1 , in particular in claims 1 to 37. Such valve is herewith incorporated into the present application.

The circumstance that channels are interconnected via a node means that the channels are in fluid communication with each other, wherein preferably the channels are connected to the node. Therein, the channels are preferably connected directly to the node. Preferably, the channels are connected to the node by their respective ends. The node is a branch node and in a certain way similar to a node in an electrical circuit. In a simplified view (e.g. ignoring compressibility of fluids) and in analogy to Kirchhoff’s first law, the sum of the fluids flowing into the node is equal to the sum of the fluids flowing out of the node.

The positioner is for positioning an object within the intermediate channel. In particular the positioner can be denoted as positioning device. Preferably, the positioner comprises a narrowing of the cross-section of the intermediate channel wherein the narrowing is preferably arranged between two sections of the intermediate channel comprising a wider cross-section. This enables an object which flows with the fluid into the intermediate channel to be positioned (i.e. trapped) by the narrowing. In an alternative preferred embodiment, the positioner comprises a widening of the crosssection of the intermediate channel wherein the widening is preferably arranged between two sections of the intermediate channel comprising a narrower cross-section. This enables an elastic object with a larger cross-section as the wider cross-sections to be squeezed through the wider cross-sections by means of the fluid. The elastic object can be positioned within the widening when it reaches the widening comprising a larger cross-section than the object. For example, this can be realized by providing a stop within the widening or by decreasing the fluid pressure when the object reaches the widening. In an alternative (however not preferred embodiment) the positioning function is fulfilled by the intermediate channel itself without an additional structure. For example, the intermediate channel might be of constant cross-section and the positioning of an elastic object with a wider cross-section that the one of the intermediate channel can be realized by squeezing the object into the channel and controlling the fluid pressure in order to move or stop the squeezed object.

In a preferred embodiment, the object which can be positioned by a positioner is a matrix, preferably a hydrogel matrix. . According to one embodiment, the matrices disclosed herein are preferably spherical, e.g. spherical hydrogel matrices but other forms may also be applied. In a preferred embodiment, the object comprises a matrix formed using droplet microfluidics. For example, a flow focusing geometry can be used for the generation of highly monodisperse droplets having a spherical shape. If the droplet diameter is larger than the width/height of the microfluidic channel in which the hydrogel formation may occur, formed matrices have a plug-like shape. In addition, matrices may be formed by conventional pipetting. Thus, matrix solutions comprising monomers, pre-polymers, precursors, polymer and/or building blocks for gelation/polymerization/curing reactions may be pipetted on a 2D surface resulting in the formation of a droplet having the shape of a spherical segment and/or a hemispherical shape. The shape depends on the surface tension between the droplet and the surrounding surfaces and may be adjusted by changing the surface characteristics. In another embodiment, matrix solutions comprising monomers, pre-polymers, precursors, polymer and/or building blocks for gelation/polymerization/curing reactions may be pipetted into a geometry having a pre-defined shape (e.g. a cylindrical geometry). Thus, matrices may assume the shape of the container containing the matrix solution during matrix formation. According to one embodiment, the object has a diameter of < 1000 pm, such as < 800 pm, < 600 pm or < 400 pm, preferably < 200 pm, such as 5 pm to 150 pm.

According to one embodiment, the object comprises a hydrogel matrix, which may be formed upon the gelation/polymerization/curing of a monomer, pre-polymer, precursor, polymer and/or building block. In one embodiment, the object, preferably having a spherical shape, comprises a hydrogel, a polymer or pre-polymer which is selected from the group comprising polyacrylamide, poly( lactic acid) (PLA), polyglycolide (PGA), copolymers of PLA and PGA (PLGA), poly(vinyl alcohol) (PVA), polyethylene glycol) (PEG), poly(ethylene oxide), poly(ethylene oxide )-co-poly( propylene oxide) block copolymers (poloxamers, meroxapols), poloxamines, polyanhydrides, polyorthoesters, poly(hydroxyl acids), polydioxanones, polycarbonates, polyaminocarbonates, poly( vinyl pyrrolidone), poly(ethyl oxazoline), carboxymethyl cellulose, hydroxyalkylated celluloses such as hydroxyethyl cellulose and methylhydroxypropyl cellulose, and natural polymers such as nucleic acids, polypeptides, polysaccharides, chitosan or carbohydrates such as polysucrose, hyaluranic acid, dextran and similar derivatives thereof, heparan sulfate, chondroitin sulfate, heparin, or alginate, and proteins including without limitation gelatin, collagen, albumin, or ovalbumin, or copolymers, or blends thereof. In particularly preferred embodiments, the monomers can be selected from polyactic acid) (PLA), poly(vinyl alcohol) (PVA), polyethylene glycol (PEG) and polyoxazoline (POx). The object preferably comprises matrix which may comprise polymers and/or precursor molecule, preferably in a predominantly crosslinked form, which have been disclosed in PCT/EP2018/074527, in particular, polymers and/or precursor molecules disclosed in claims 101 to 155, which are herein incorporated by reference.

In a preferred embodiment, the object comprises a hydrogel. The hydrogel may be a hydrogel as disclosed in PCT/EP2018/074527, in particular, hydrogels as disclosed in claims 1 to 51 and 72, which are herein incorporated by reference. PCT/EP2018/074527 further discloses methods for producing a hydrogel in claims 52 to 71 , which are herein incorporated by reference. Furthermore, a kit for producing a hydrogel is disclosed in PCT/EP2018/074527 in claims 99 and 100, which are also herein incorporated by reference.

According to a preferred embodiment, the object comprises a hydrogel matrix comprising one or more selected from the group consisting of a virus particle, a vesicle, such as an exosome or an apoptotic vesicle, a cell, such as prokaryotic cell, e.g. bacteria, or a eukaryotic cell and/or a combination thereof. Accordingly, the hydrogel matrix may contain one or more types and/or sizes of compounds of said group. For instance, the hydrogel matrix may comprise a cell, such as a eukaryotic cell and a virus. Or the hydrogel matrix may comprise a vesicle and a cell. According to a preferred embodiment, the object comprises at least one cell. Optionally, the object is a cell. According to a preferred embodiment, the object comprises a hydrogel matrix comprising at least one cell, also referred to as cell-laden hydrogel matrix. According to a one embodiment, the object comprises a hydrogel matrix that provides a three-dimensional environment to the at least one cell, wherein preferably the matrix is at least 5 pm and < 200 pm in diameter.

According to a preferred embodiment, the object comprises a hydrogel matrix, preferably having a spherical shape, comprising more than one cell. According to another embodiment, the hydrogel matrix comprises a colony of cells. Preferably, a colony of cells can be located inside the three-dimensional matrix. According to another embodiment, the cell number changes throughout performing the method. For instance, the cell number increases over the course of cultivation, decreases over the course of cultivation or remains constant over the course of cultivation. A colony of cells may be formed by proliferation of one or more cells, wherein preferably cells proliferate inside the three-dimensional matrix. In another embodiment, the hydrogel matrix comprises at least two different types of cells that interact. In particular, the hydrogel matrix may comprise two different types of cells that interact.

The at least one cell may be selected from a prokaryotic and/or an eukaryotic cell. The at least one cell may be selected from the groups consisting of bacteria, archaea, plants, animals, fungi, slime moulds, protozoa, and algae. According to a preferred embodiment, the at least one cell may be selected from animal cells, preferably human cells. According to one embodiment, the at least one cell may be selected from cell culture cell lines. According to another embodiment, the at least one cells may be selected from the group consisting of stem cells, bone cells, blood cells, muscle cells, fat cells, skin cells, nerve cells, endothelial cells, sex cells, pancreatic cells, and cancer cells. According to another embodiment, the at least one cell may be derived from cells of the nervous system, the immune system, the urinary system, the respiratory system, the hepatopancreatic-biliary system, the gastrointestinal system, the skin system, the cardiovascular system, developmental biology (including stem cells), pediatrics, organoids, and model organisms. According to another embodiment, the at least one cell may be derived from one or more of blood and immune system cells, including erythrocytes, megakaryocytes, platelets, monocytes, connective tissue macrophages, epidermal Langerhans cells, osteoclast (in bone), dendritic cells, microglial cells, neutrophil granulocytes, eosinophil granulocytes, basophil granulocytes, hybridoma cells, mast cells, helper T cells, suppressor T cells, cytotoxic T cells, natural killer T cells, B cells, natural killer cells, reticulocytes, hematopoietic stem cells, and committed progenitors for the blood and immune system.

Preferably, the second (forth) channel is connected to the inlet node with his first end and connected to the first (second) intermediate node with his second end. Thus, the second channel extends from the inlet node to the first intermediate node and/or the forth channel extends from the inlet node to the second intermediate node. Preferably, they do not extend beyond according nodes. Preferably, the first (third) channel is connected to the first (second) intermediate node with his first end and connected to the outlet node with his second end. Thus, the first channel extends from the first intermediate node to the outlet node and/or the third channel extends from the second intermediate node to the outlet node. Preferably, they do not extend beyond the according nodes. Preferably, the intermediate channel is connected to the first intermediate node with his first end and connected to the second intermediate node with his second end. Thus, the intermediate channel extends from the first intermediate node to the second intermediate node. Preferably, it does not extend beyond the first and second intermediate node.

In a preferred embodiment the resistances, channels and valves are configured in such a way that no fluid can flow through the intermediate channel if the valves are opened. In analogy to the Wheatstone bridge in the field of electrical engineering, this state can be achieved by fulfilling the equation R1/R2 = R3/R4.

In a preferred embodiment the resistances, channels and valves are configured in such a way that a fluid can flow through the intermediate channel from the first intermediate node to the second intermediate node if the valves are opened. Thereby, the fluid flows mainly from the second channel to the third channel via the intermediate channel. This has the advantage that an object entered the intermediate node form the first intermediate node and then being trapped within the positioner remains safely within the positioner due to the fluid flow pushing it into the positioner.

In a preferred embodiment the resistances, channels and valves are configured in such a way that a fluid can flow through the intermediate channel from the first intermediate node to the second intermediate node if the first valve is opened and the second valve is closed. Thereby, the fluid flows mainly from the second channel to the third channel via the intermediate channel.

In a preferred embodiment the resistances, channels and valves are configured in such a way that a fluid can flow through the intermediate channel from the second intermediate node to the first intermediate node if the second valve is opened and the first valve is closed. Thereby, the fluid flows mainly from the fourth channel to the first channel via the intermediate channel. This fluid flow allows releasing an object, which entered the intermediate channel form the first intermediate node and is trapped within the positioner, from the positioner and transport it by the fluid flow to the outlet channel via the first channel.

In a preferred embodiment the resistances, channels and valves are configured in such a way that no fluid can flow through the intermediate channel if the valves are completely closed. In one embodiment R1 and R3 are equal or deviate from each other by at most 5%. Additionally, or alternatively, R2 and R4 are equal or deviate from each other by at most 5%. In a preferred embodiment R1, R2, R3 and R4 are equal or at least deviate from each other by at most 5%.

In one embodiment the first channel and the third channel have the equal geometry. Additionally or alternatively the second channel and the fourth channel have the equal geometry. In a preferred embodiment the first to fourth channel have the same geometry. In particular, the term “same (or equal) geometry” means the same size of cross-section, the same shape of cross-section and the same length of channel. The shape of cross-section is preferably rectangular. Alternative shapes are conceivable like circular for example. Preferably, the first and the third channel have the same cross-section (area) and/or cross-section shape. Preferably, the second and the fourth channel have the same cross-section (area) and/or cross-section shape. Particularly preferred, all these channels have the same cross-section (area) and/or cross-section shape.

The invention also refers to a system comprising a first device according to the invention and at least a second device according to the invention, wherein the devices are interconnected. In a preferred embodiment the system comprises several devices according to the invention. In particular, the devices are interconnected in such a way that they form an array.

In a preferred embodiment, the system further comprising a feeding line which is formed by a feeding inlet channel, the intermediate channel of the first device, at least one connection channel, the intermediate channel of the at least second device and a feeding outlet channel; wherein the feeding inlet channel, the first channel, the second channel and the intermediate channel are interconnected via the first intermediate node; wherein the connection channel, the third channel, the fourth channel and the intermediate channel are interconnected via the second intermediate node and the connection channel, the first channel of the second device, the second channel of the second device and the intermediate channel of the second device are interconnected via the first intermediate node of the second device; wherein the feeding outlet channel, the third channel of the second device, the fourth channel of the second device and the intermediate channel of the second device are interconnected via the second intermediate node of the second device; wherein the system further comprising a feeding inlet valve arranged at the feeding inlet channel and configured to be actuated in order to control the flow rate through the feeding inlet channel; wherein the system further comprising a feeding outlet valve arranged at the feeding outlet channel and configured to be actuated in order to control the flow rate through the feeding outlet channel; wherein the system further comprises a connection valve arranged at the connection channel and configured to be actuated in order to control the flow rate through the connection channel. This embodiment comprises the preferred case in which more than two devices are interconnected. (The system comprises “at least” a second device.) The interconnected devices form an array. For example, if the system comprises three devices, the second device and the third device are interconnected according to interconnection of the first and the second device. The feeding line is additionally formed by a second connection channel wherein the second connection channel, the third channel of the second device, the fourth channel of the second device and the intermediate channel of the second device are interconnected via the second intermediate node of the second device and the second connection channel, the first channel of the third device, the second channel of the third device and the intermediate channel of the third device are interconnected via the first intermediate node of the third device. The feeding outlet channel, the third channel of the third device, the fourth channel of the third device and the intermediate channel of the third device are interconnected via the second intermediate node of the third device. In particular, the system further comprises a second connection valve arranged at the second connection channel and configured to be actuated in order to control the flow rate through the second connection channel.

In particular, by completely closing a connection valve it is possible to disconnect one device from the other as is described in more detail below. This enables to control the fluid flow within the individual devices independently from each other.

Preferably, the connection channel is connected to the second intermediate node (of the first device) with his first end and connected to the first intermediate node of the second device with his second end. The same applies mutatis mutandis to an additional connection channel.

In a preferred embodiment the system preferably comprises a first outer valve configured to be actuated in order to control the flow rate through the inlet channel of the first device and the inlet channel of the second device, thereby preferably applying the same pressure to each inlet channel; further comprising a second outer valve configured to be actuated in order to control the flow rate through the outlet channel of the first device and the outlet channel of the second device, thereby preferably applying the same pressure to each outlet channel. This has the advantage that by activating one single valve, the flow rate through several channels can be controlled simultaneously. In particular, by generating a pressure difference between the inlet channels by means of the first outer valve and the outlet channels by means of the second outer valve a fluid flow can be induced through several devices simultaneously.

In an alternative, however also preferred embodiment, the first outer valve is not provided and/or all first and second valves are actuated in order to control or block the flow control through the appropriate inlet channels.

In a preferred embodiment the array formed by the interconnected devices comprises n times m devices, wherein the devices are arranged in such a way that they from a matrix of m columns and n rows, wherein m is a number, preferably 16, and n is a number, preferably 24, wherein the system is preferably configured in such a way that all first valves of the devices arranged in the same column can be actuated simultaneously and all first valves arranged in the same row can be actuated simultaneously and/or all second valves of the devices arranged in the same column can be actuated simultaneously and all second valves arranged in the same row can be actuated simultaneously.The last device of a column and the first device of the next column are interconnected like the first device and the second device of the same column for example. However, if necessary, a longer connection channel might be used. This preferred embodiment allows choosing easily the valves of the device intended to be operated.

A 16x24-matrix is suitable to be integrated into the footprint of a conventional glass slide (26 x 76 mm) for example. Preferably, the footprint of such matrix is 50 mm times 16 mm (= 800 mm²). In a further preferred embodiment, n is 32 and m is also 32 so that in total there are 1024 devices. In a further preferred embodiment, m is 16 and n is 8. Preferably, the footprint of such matrix is 15 mm times 17 mm (= 255 mm²).

The invention also refers to a method comprising the steps: providing a system according to the invention with the first valve and the second valve of the first device are closed, the first valve and the second valve of the second device are closed, the second outer valve is closed, whereas the feeding inlet valve, the connection valve and the feeding outlet valve are opened; causing a fluid flow within the feeding line from the feeding inlet channel to the feeding outlet channel; providing an object within the feeding inlet channel; transporting the object from the feeding inlet channel within the feeding line by the flow of the fluid.

In particular, a fluid flow within the feeding line from the feeding inlet channel to the feeding outlet channel is caused by a pressure difference between the feeding inlet channel and the feeding outlet channel. The object is for example a cell or particle or a cell-laden spherical hydrogel matrix. In particular, the object flows into the intermediate channel and is stopped in the positioner and preferably trapped by its narrowing or widening of the cross-section.

In a preferred embodiment the object is trapped by the positioner and the method comprises the further steps: providing a second object within the feeding inlet channel; transporting the object from the feeding inlet channel to the positioner of the second device, wherein the positioner of the first device comprises a structure allowing the second object passing it while the first object is trapped, the structure being preferably a bypass. For example, if a first object is trapped by the positioner by the narrowing of the cross-section, the second object would be led through the bypass allowing the second object to continue its flow to the intermediate channel of the second device in order to be trapped in the positioner of the second device. Therewith, it is possible to “feed” the positioners of all devices of an array with an object by means of a single feeding line. In a preferred embodiment, the first channel and the third channel can be used as bypass. If the positioner is occupied by a first particle, the flow resistance in the intermediate channel rises and a second particle entering the device through the feeding inlet node can flow through the first channel and third channel due to a less resistance value of this path. In particular, this can be achieved by closing the first valve and the second valve as well as closing the outlet valve.

The invention also refers to a method comprising the steps: providing a system according to the invention with the feeding inlet valve, the (or each) connection valve and the feeding outlet valve are closed, the second outer valve is opened, comprising the following steps: actuating the first valve of all devices of column x simultaneously, wherein x is a (natural) number of a range from 1 to m, and/or actuating the second valve of all devices of row y simultaneously, wherein y is a (natural) number of a range from 1 to n in order to control the flow rate and flow direction of a fluid within the intermediate channel of the device which is located in column x and row y. Thereby, a more flexible operation of the system is possible: In particular, the flow rate and flow direction within the intermediate channel of one device can be controlled independently of other devices.

In a preferred embodiment, the method comprising the steps: providing a system according to the invention with the feeding inlet valve, the (or each) connection valve and the feeding outlet valve are closed, the first valve of all devices is opened, the second valve of all devices is opened, comprising the following steps: Closing the second valve of all devices, except of the devices in column x, and closing the first valve of all devices in row y, thereby causing a flow from the second intermediate node to the first intermediate node within the intermediate channel of the device which is located in column x and row y. In particular, this device is the only device with such flow direction within the intermediate channel. In particular, this flow direction is used to transfer an object which is located within the positioner of the intermediate channel in a common outer channel or to another position of the system. In a further preferred embodiment, the method additionally comprises the step of closing the first valve of all devices in row y2, thereby causing a flow from the second intermediate node to the first intermediate node within the intermediate channel of the device which is located in column x and row y2, wherein row y2 is a different row than row y. This embodiment enables to transfer an object which is located within the positioner of the intermediate channel of the device which is located in column x and row y2 in a common outer channel or to another position of the system. It is preferred to close the first valve of all devices in row y and the first valve of all devices in row y2 simultaneously. Thereby, it is possible to transfer several objects simultaneously.

In a preferred embodiment, the method comprising the steps: providing a system according to the invention with the feeding inlet valve, the (or each) connection valve and the feeding outlet valve are closed, the first valve of all devices is opened, the second valve of all devices is closed, comprising the following steps: Opening the second valve of all devices in column x and closing the first valve of all devices in row y, thereby causing a flow from the second intermediate node to the first intermediate node within the intermediate channel of the device which is located in column x and row y. In particular, this device is the only device with such flow direction within the intermediate channel. In particular, this flow direction is used to transfer an object which is located within the positioner of the intermediate channel in a common outer channel or to another position of the system.

The device and system according to the invention can be preferably used for carrying out biotechnological processes. One example is an incubation process. For example the device and system can be used for treatment and forming of objects like cells (e.g. human or plant cells) and particles. In particular, the system can be used for the cultivation of single or multiple cells of the same or of different type located within spherical hydrogel matrices.

The invention is described by the following figures wherein the figures only illustrate exemplary embodiments of the invention. In particular, the subject matter of the invention is not limited to these embodiments.

Figure 1A shows an embodiment of the device according to the invention.

Figure 1 B shows a modification of the embodiment according to figure 1 A.

Figure 2 shows an embodiment of the system according to the invention.

Figure 2A shows the system according to figure 2 with a different valve configuration.

Figures 3A, 3B and 3C show an exemplary embodiment of the device according to the invention.

Figure 4 is an exemplary system according to the invention.

Figures 5A to 5C show an exemplary embodiment as 4x4-matrix with different valve configurations.

Figure 6 shows a system consisting of three 8x16-matrixes. FPX: Foot print in X direction. FPY: Foot print in Y direction.

Figure 1 shows an embodiment of the device 10 according to the invention. The device 10 is a microfabricated device. It comprises an inlet channel IC, outlet channel OC, first channel C1 having a hydrodynamic resistance R1 , second channel C2 having a hydrodynamic resistance R2, third channel C3 having a hydrodynamic resistance R3, fourth channel C4 having a hydrodynamic resistance R4 and a intermediate channel 0 having a hydrodynamic resistance R0. The intermediate channel comprises a positioner. The positioner is preferably a widening but could also be a narrowing of the cross section of the intermediate channel 0.

The device 10 further comprises an inlet node IN, outlet node ON, first intermediate node IN1 and second intermediate node IN2.

The inlet channel IC, second channel C2 and fourth channel C4 are interconnected via the inlet node IN. The outlet channel OC, first channel C1 and third channel C3 are interconnected via the outlet node ON. The first channel C1 , second channel C2 and intermediate channel 0 are interconnected via the first intermediate node IN1. The third channel 03, fourth channel 04 and intermediate channel 0 are interconnected via the second intermediate node IN2.

The device 10 comprises a first valve V1 arranged at the second channel 02 and configured to be actuated in order to control the flow rate through the second channel 02. In contrast to known devices, the device according to figure 1 comprises a second valve V2 arranged at the fourth channel 04 and configured to be actuated in order to control the flow rate through the fourth channel 04. Thereby, it is possible to control the flow rate of a fluid flowing through the device 10 as flexibly as desired but without or less geometric constraints for designing the channels. All fluid flow configurations can be realized in an easy way by actuating the first valve V1 and/or the second valve V2. For example R1 to R4 may have the same geometry. The skilled person knows how to choose the different resistances of the channels in combination with the actuation of the valves in order to create a specific flow rate. For example, the analogy of the Wheatstone bridge might be helpful. With reference to the present invention, the following formula of the Wheatstone bridge might be considered:

UO = Ux((R1xR4)-(R2xR3))/((R1+R2)x(R3+R4)), wherein

UO is the pressure difference between the first intermediate node IN1 and second intermediate node IN2 and II is the pressure difference between input node IN and output node ON.

By actuating the first valve V1, it is in principle possible to change the resistance for a fluid flowing through the first channel C1. By actuating the second valve V2, it is in principle possible to change the resistance for a fluid flowing through the fourth channel C4. Therefore, by defining the channels with certain resistances and actuating the valves, it is possible to determine UO. For example, if UO is zero, no fluid flows through the intermediate channel. For example, if UO is positive, a flow from the first intermediate node IN1 to the second intermediate node IN2 takes place, whereas a negative value represents a flow in the opposite direction.

Other than in the known devices, it is in particular not necessary to define the first channel C1 and the third channel C3 with a different geometry, in particular with a different length. In principle, it is even possible to define the channels C1 to C4 with the same geometry and nevertheless achieve a fluid flow as desired. However, it is preferred to design the channels differently.

Figure 1B shows the embodiment of figure 1 comprising additionally a feeding inlet channel FIC having a feeding inlet valve which is described in detail below.

Figure 2 shows an embodiment of the system according to the invention. It comprises a first device 10, second device 20, third device 30 and fourth device 40 according to the invention, wherein the devices are interconnected. The system further comprises a feeding line FL formed by a feeding inlet channel FIC, the intermediate channel of all devices and a first connection channel CC1, a second connection channel CC2 and a third connection channel CC3 as well as a feeding outlet channel FOC. This exemplary embodiment is an array of four devices which are interconnected by the interconnection channels as described above. Moreover, the system comprises a feeding inlet valve FIV arranged at the feeding inlet channel FIC and configured to be actuated in order to control the flow rate through the feeding inlet channel FIC. The system further comprises a feeding outlet valve FOV arranged at the feeding outlet channel FOC and configured to be actuated in order to control the flow rate through the feeding outlet channel FOC. The system further comprises a connection valve CC1-V arranged at the first connection channel, a second connection valve CC2-V arranged at the second connection channel and a third connection valve CC3-V arranged at the third connection channel. The embodiment of figure 2 illustrates a 2x2-matrix, where the first and second device are arranged in the first column and the third and fourth device are arranged in the second column. The first line is formed by the first and third device, whereas the second line is formed by the second and fourth device.

Figure 2 shows also a first outer valve OV1 configured to be actuated in order to control the flow rate through the inlet channels of the first to fourth device 10 to 40. A second outer valve OV2 is also provide and configured to be actuated in order to control the flow rate through the outlet channels of the first to fourth device 10 to 40. The valves in the embodiment according to figure 2 are all opened.

In figure 2A, the system according to figure 2 is shown in such a state that all first and second valves of the devices and the second outer valve OV2 are completely closed. Closed valves are marked by dotted rectangles. Thus, a fluid flow from the inlet channels to the outlet channels is not possible. Moreover, the valves of the fluid line are opened, i.e. the valves FIV, CC1-V to CC3-V and FOV. This configuration has the advantage that a fluid can flow through the feeding line from the feeding inlet channel FIC to the feeding outlet channel FOC. Thereby, it is possible to provide objects within the feeding inlet channel FIC and provide them to the individual positioners sequentially. Figure 2B shows the system according to figure 2 with a different valve-actuation configuration. All valves of the feeding line are completely closed. Thereby, the different devices are disconnected from each other. This has the advantage that the fluid flow of each device can be generated individually.

For example, in the first device 10 a fluid flow is established from the fourth channel 4 to the first channel 1 via the intermediate channel. The fluid flows through the intermediate channel from the second intermediate node to the first intermediate node. This is possible since the second valve V2 is opened and the first valve V1 is closed. Therefore, it is possible to release an object which is trapped in the positioner of the intermediate channel.

The second device 20 shows a different actuation of valve V1 and valve V2. Here, both valves are closed whereby no fluid flows through the intermediate channel.

The third device 30 comprises two opened valves V1 and V2. The fluid flows mainly form the second channel to the third channel via the intermediate channel. The fluid flows through the intermediate channel from the first intermediate node to the second intermediate node.

The first valve V1 of the fourth device 40 is opened, whereas the second valve V2 is closed. The fluid flows mainly form the second channel to the third channel via the intermediate channel. The fluid flows through the intermediate channel from the first intermediate node to the second intermediate node.

Figures 3A, 3B and 3C show an exemplary embodiment of the device according to the invention. The reference signs in this figures which are the same as the reference signs in figures 1 A, 1 B and 2 refer to the same features as in this figures.

As shown in figure 3A, the inlet channel IC is a vertical channel and the arrow indicates the direction of a fluid flowing into the inlet channel. The outlet channel OC has a vertical section at his firs end and the arrow indicates the direction of a fluid flowing out of the outlet channel. The second end of the outlet channel OC is connected to the outlet node ON. Furthermore, an exemplary embodiment of the positioner P is shown. The positioner P is arranged within the intermediate channel and forms a widening of this channel. Within the positioner P there is a trapping structure configured to trap a particle which flows into the intermediate channel through the first intermediate node FIN. On each side of the trapping structure a bypass channel BC is arranged to enable a second particle to pass the positioner P and flow to the second intermediate node IN2 if a first particle is already positioned within the trapping structure of the positioner P. The feeding inlet channel FIC has a feeding inlet valve FIV which is a membrane and can be deformed in order to control the flow rate through the feeding inlet channel FIC. Furthermore, the first valve V1 and the second valve V2 are elastic elements which can be deformed to control the flow rate through the second channel C2 and the fourth channel C4. The valves V1 and V2 are equal in shape and size. The fourth channel comprises two sections. The first section is the valve V2 and the second section is the part between the inlet node IN and the valve V2. The first channel C1 is formed by the first valve V2. The height and width of the intermediate channel, the first channel C1 and the outlet channel OC is almost the same. These channels are provided to transport a particle of e.g. 80 pm. Compared with the other channels (e.g. third channel C3) these channels have a larger cross-section. In this embodiment, the first channel C1 and the third channel C3 have the same resistance. Due to the fact that the first channel has a larger cross-section, it must be longer to cause the same pressure loss.

Figure 3B shows the device of figure 3A in the top view. Figure 3C depicts the device of figures 3B with a schematically indication of the resistances of the channels C1 to C4. The first channel C1 has the resistance R1 and the third channel C3 has the resistance R3. The second channel C2 has the total resistance R2* which is the sum of the resistance R2 of channel C2 and the resistance RiV1 of the first valve V1 in the opened stated. The resistance of the first valve RiV1 can be changed by closing the valve. Therefore, RiV1 is a flexible resistance. The same applies mutatis mutandis to total resistance R4* which is the sum of the resistance R4 of the fourth channel C4 and the resistance of the second valve RiV2. Furthermore, RiV3 is illustrated which is the flexible resistance of the feeding inlet valve FIV.

Figure 4 is an exemplary system according to the invention. In principle it is an array as depicted in schematically diagram of figure 2. However, the connection channels connecting the devices 10 to 40 are not illustrated for a better overview. Further, a common channel COC connecting the outlet channels of the individual devices is shown.

Figures 5A to 5C show an exemplary embodiment as 4x4-matrix with different valve configurations. The same pressure must be applied at each inlet. This is guaranteed by the tree structure of the supply channel. Figure 6 shows a system consisting of three 8x16-matrixes.

It is preferred that (all) valves V1 of one column may be operated simultaneously using one external (solenoid) valve. Additionally or alternatively, (all) valves V2 of one row may be operated simultaneously using on (external) solenoid valve. Thus, for an exemplary 4x4-matrix (four columns and four rows), eight external valves are needed. For an exemplary 16x24-matrix (16 columns and 24 rows), 40 external valves are needed.

Figure 6 shows an illustration of an exemplary system having 16 x 24 devices. The length in X direction (FPX) is 50 mm and the length in Y direction (FPY) is 16 mm resulting in a footprint of 800 mm² in total. Thus, the mean footprint of one device is 2.08 mm².

Claims

Claims Device (10) comprising an inlet channel (IC), outlet channel (OC), first channel (C1) having a hydrodynamic resistance R1, second channel (C2) having a hydrodynamic resistance R2, third channel (C3) having a hydrodynamic resistance R3, fourth channel (C4) having a hydrodynamic resistance R4, intermediate channel (0) having a hydrodynamic resistance R0 and a positioner (P) for positioning an object within the intermediate channel (0), the positioner (P) preferably comprising a narrowing of the cross section of the intermediate channel (0); the device (10) further comprising an inlet node (IN), outlet node (ON), first intermediate node (IN1) and second intermediate node (IN2); wherein the inlet channel (IC), second channel (C2) and fourth channel (C4) are interconnected via the inlet node (IN), the outlet channel (OC), first channel (C1) and third channel (C3) are interconnected via the outlet node (ON), the first channel (C1), second channel (C2) and intermediate channel (0) are interconnected via the first intermediate node (IN1) and the third channel (C3), fourth channel (C4) and intermediate channel (0) are interconnected via the second intermediate node (IN2); the device (10) further comprising a first valve (V1) arranged at the second channel (C2) and configured to be actuated in order to control the flow rate through the second channel (C2), characterized in that the device (10) comprises a second valve (V2) arranged at the fourth channel (C4) and configured to be actuated in order to control the flow rate through the fourth channel (C4). Device (10) according to the previous claim, wherein the resistances, channels and valves are configured in such a way that no fluid can flow through the intermediate channel (0) if the valves (V1, V2) are opened or alternatively the resistances, channels and valves are configured in such a way that a fluid flows through the intermediate channel (0) from the first intermediate (IN1) node to the second intermediate node (IN2) if the valves (V1, V2) are opened. Device (10) according to one of the previous claims, wherein the resistances, channels and valves are configured in such a way that a fluid can flow through the intermediate channel (0) from the first intermediate node (IN1) to the second intermediate node (IN2) if the first valve (V1) is opened and the second valve (V2) is closed. Device (10) according to one of the previous claims, wherein the resistances, channels and valves are configured in such a way that a fluid can flow through the intermediate channel (0) from the second intermediate node (IN2) to the first intermediate node (I N 1 ) if the second valve (V2) is opened and the first valve (V1) is closed. Device (10) according to one of the previous claims, wherein R1 and R3 are equal or deviate from each other by at most 5% and/or wherein R2 and R4 are equal or deviate from each other by at most 5%.

6. Device (10) according to one of the previous claims, wherein the first channel (C1) and the third channel (C3) have the equal geometry and/or wherein the second channel (C2) and the fourth channel (C4) have the equal geometry.

7. System comprising a first device (10) according to one of the previous claims and at least a second device (20) according to one of the previous claims, wherein the devices (10, 20) are interconnected.

8. System according to the previous claim, further comprising a feeding line (FL) which is formed by a feeding inlet channel (FIC), the intermediate channel (0) of the first device (C1), at least one connection channel (CC1), the intermediate channel (0) of the at least second device (20) and a feeding outlet channel (FOC); wherein the feeding inlet channel (FIC), the first channel (C1), the second channel (C2) and the intermediate channel (0) are interconnected via the first intermediate node (IN1); wherein the connection channel (CC1), the third channel (C3), the fourth channel (C4) and the intermediate channel (0) are interconnected via the second intermediate node (IN2) and the connection channel (CC1), the first channel (C1) of the second device (20), the second channel (C2) of the second device (20) and the intermediate channel (0) of the second device (20) are interconnected via the first intermediate node (I N 1) of the second device (20); wherein the feeding outlet channel (FOC), the third channel (C3) of the second device (20), the fourth channel (C4) of the second device (20) and the intermediate channel (0) of the second device (20) are interconnected via the second intermediate node (IN2) of the second device (20); wherein the system further comprising a feeding inlet valve (FIV) arranged at the feeding inlet channel (FIC) and configured to be actuated in order to control the flow rate through the feeding inlet channel (FIC); wherein the system further comprising a feeding outlet valve (FOV) arranged at the feeding outlet channel (FOC) and configured to be actuated in order to control the flow rate through the feeding outlet channel (FOC); wherein the system further comprises a connection valve (CC1-V) arranged at the connection channel (CC1) and configured to be actuated in order to control the flow rate through the connection channel (CC1).

9. System according to the previous claim, further preferably comprising a first outer valve (OV1) configured to be actuated in order to control the flow rate through the inlet channel (IC) of the first device (10) and the inlet channel (IC) of the second device (20), thereby preferably applying the same pressure to each inlet channel (IC); further comprising a second outer valve (OV2) configured to be actuated in order to control the flow rate through the outlet channel (OC) of the first device (10) and the outlet channel (OC) of the second device (20), thereby preferably applying the same pressure to each outlet channel.

10. System according to one of the previous system claims, wherein the array formed by the interconnected devices comprises n times m devices, wherein the devices are arranged in such a way that they form a matrix of m columns and n rows, wherein m 19 is a number, preferably 16, and n is a number, preferably 24, wherein the system is preferably configured in such a way that all first valves (V1) of the devices arranged in the same column can be actuated simultaneously and all first valves (V1) of the devices arranged in the same row can be actuated simultaneously and/or all second valves (V2) of the devices arranged in the same column can be actuated simultaneously and all second valves (V2) of the devices arranged in the same row can be actuated simultaneously. Method comprising the steps: providing a system according to one of the previous system claims with the first valve (V1) and the second valve (V2) of the first device are closed, the first valve (V1) and the second valve (V2) of the second device are closed, the second outer (OV2) valve is closed, whereas the feeding inlet valve (Fl V) , the connection valve (CC1) and the feeding outlet valve (FOV) are opened; causing a fluid flow within the feeding line (FL) from the feeding inlet channel (FIC) to the feeding outlet channel (FOC); providing an object within the feeding inlet channel (FIC); transporting the object from the feeding inlet channel (FIC) within the feeding line by the flow of the fluid. Method according to the previous claim, wherein the object is trapped by the positioner of the first device (10); further comprising the steps: providing a second object within the feeding inlet channel (FIC); transporting the object from the feeding inlet channel (FIC) to the positioner (P) of the second device (20), wherein the positioner (P) of the first device (10) comprises a structure allowing the second object passing it, the structure being preferably a bypass. Method according to the previous claims, wherein the first and/or second object has one or more of the following characteristics: the object comprises a matrix, in particular a hydrogel matrix, and preferably has a spherical shape; the object comprises a matrix, preferably a hydrogel matrix, formed using droplet microfluidics; the object comprise a hydrogel matrix formed upon the gelation/polymerization/curing of a monomer, pre-polymer, precursor, polymer and/or building block; the object comprises at least one cell, optionally the object is a cell; and/or the object comprises a hydrogel matrix comprising at least one cell, optionally, o the hydrogel matrix comprises one cells; o the hydrogel matrix comprises a colony of cells of the same type; o the hydrogel matrix comprises two or more different types of cells; or o the hydrogel matrix comprises colonies of two or more different types of cells. Method comprising the steps: providing a system according to one of the previous system claims with the feeding inlet valve (FIV), the connection valve (CC1-V) and 20 the feeding outlet valve (FOV) are closed, the second outer valve (OV2) is opened, comprising the following steps: actuating the first valve (V1) of all devices of column x simultaneously, wherein x is a number of a range from 1 to m, and/or actuating the second valve (V2) of all devices of row y simultaneously, wherein y is a number of a range from 1 to n in order to control the flow rate and flow direction of a fluid within the intermediate channel (0) of the device which is located in column x and row y.