Cell migration study device

Abstract

The present invention relates to a device for studying cell migration, which includes a first chamber and a second chamber with a substrate in between that has a set of through-holes for fluidic connection. The through-holes come in at least two types with different sizes and / or shapes. A delivery system is adapted to create both uniform and gradient fields of a chemical substance across the chambers, facilitating the study of cell migration in response to chemical stimuli. The device may include detection means for monitoring cell migration and can be used with various cell types, including those that mimic the blood-brain barrier. The substrate may be made of silicon and have through-holes of various shapes, and the device can be optically transparent for imaging purposes. The delivery system can include a microfluidic system with adjustable gradient steepness, and the device may be integrated with a data processing unit for analyzing migration data.

Description

[0001] Cell Migration Study Device

[0002] Field of the Invention

[0003] The present invention relates to the field of biomedical devices, and more specifically to a device designed for the study of cellular behaviors such as migration, transmigration, and pathogen translocation.

[0004] Background of the Invention

[0005] The study of cellular behavior, particularly cell migration, is a critical area of research in the field of cell biology and medicine. Cell migration is a fundamental process that underlies many physiological and pathological events, including embryonic development, wound healing, immune responses, and the progression of diseases such as cancer and inflammation. Transmigration, a specific type of cell migration, involves the movement of cells of the body (e.g., immune cells or tumor cells) across a barrier, such as the endothelium, which lines the interior surface of blood vessels. This process is essential for the immune system to function correctly, allowing immune cells to reach sites of infection or injury. Another type of migration: bacterial or pathogen translocation, involves migration of pathogens or bacteria across a cell barrier, e.g. the intestinal barrier, to enter the systemic circulation and gain access to distant organs.

[0006] One of the most significant barriers in the human body is the blood-brain barrier (BBB), which serves as a protective shield for the central nervous system (CNS). The BBB regulates the passage of substances between the bloodstream and the brain, preventing harmful agents from entering the delicate neural environment. Similarly, other barriers like the intestinal barrier play a crucial role in maintaining homeostasis and preventing the translocation of pathogenic bacteria into the systemic circulation.

[0007] Understanding the mechanisms of cell transmigration and pathogen translocation is vital for developing new therapeutic strategies to treat various diseases. However, studying these processes presents several challenges. Current testing systems often focus on a limited number of parameters, such as attractant type and concentration, and typically rely on a single counting mechanism, like fluorescence microscopy. This approach may not fully capture the complexity of cell transmigration or pathogen translocation as these processes are influenced by multiple factors.

[0008] Moreover, most existing testing setups observe migrating cells by monitoring the barrier, with some systems using changes in electrical properties across the entire barrier. While these methods have provided valuable insights, they may not offer the flexibility needed to study cellular transmigration and translocation as they occur in nature. There is a need for a more comprehensive and versatile testing platform that can simultaneously monitor multiple parameters and their potential interdependence, allowing for a more accurate representation of biological scenarios.

[0009] Despite the progress made in the field, there remains a need for further advancements in the development of testing platforms that can provide a more detailed and integrated understanding of cell transmigration and pathogen translocation. These advancements would significantly contribute to the study of physiological and pathological processes in humans, potentially leading to breakthroughs in medical treatments and diagnostics.

[0010] Summary of the Invention

[0011] It is an object of embodiments of the present invention to enable the study of cellular migration, e.g., transmigration or translocation across various barriers, with a high degree of flexibility in monitoring multiple influencing parameters simultaneously. This objective is accomplished by a device for studying cell migration according to the invention.

[0012] In the first aspect, the present invention relates to a device for studying cell migration, comprising a first chamber, a second chamber having a first and a second sides, a substrate delimiting the first chamber from the second chamber and comprising a set of through-holes for fluidically connecting the first chamber and the second chamber, said set comprising at least two types of through-holes differing in their geometries, and a delivery system adapted for delivering a chemical substance from the first and the second sides, thereby enabling the establishment of both uniform fields and gradient fields of said chemical substance between the first side and the second side in a first direction parallel to the substrate.

[0013] In embodiments, the first and second sides may be opposite to one another. This embodiment provides the advantage of a clear and straightforward design for the establishment of chemical gradients.

[0014] In embodiments, two chemical gradients with different chemicals may be created in the second chamber. For instance, one gradient can be in an opposed, but parallel, direction to the other. In other embodiments, the second chamber may further comprise a third and a fourth side, and the delivery system may be adapted for delivering a chemical substance from the third and the fourth sides thereby enabling the establishment of a gradient field of the second chemical substance between the third side and the fourth side in another direction parallel to the substrate. This other direction may be perpendicular to the first direction, but not exclusively. This embodiment allows for the creation of more complex chemical environments within the device.

[0015] In embodiments, the device may further comprise detection means for detecting the migration of cells through each through-hole. This embodiment offers the advantage of enabling the monitoring and quantification of cell migration events.

[0016] In embodiments, the geometry of each through-hole may be determined at the top surface of the substrate, i.e., the side of the substrate facing the first chamber.

[0017] Preferably, the geometry of each through-hole may be constant or substantially constant across the thickness of the substrate. In embodiments, each through-hole has a cross-section not varying by more than 10%, preferably not varying by more than 5%, around an average cross-section across the thickness of the substrate.

[0018] In embodiments, the set of through-holes may be organized so that the through- holes are aligned in parallel rows and lines.

[0019] In embodiments, any through-hole may be misaligned with respect to its neighboring through-holes in a line or a column by no more than 500 nm. This level of precision is achievable through lithographic manufacturing techniques.

[0020] In embodiments, the thickness of the substrate may range from 1 pm to 100 pm, preferably from 5 to 50 pm, more preferably from 10 pm to 50 pm. This thickness range is specific to silicon substrates and cannot easily be achieved with polymer-based materials while keeping similar mechanical properties. In embodiments, said set of through-holes may be organized such that, along at least a second direction parallel to the substrate, each successive through-hole is of equal or larger cross-sectional area relative to an immediately preceding through-hole if present, culminating in a final through-hole having a cross-sectional area greater than that of at least another through- hole in that direction. This embodiment allows for the study of cell migration across a range of through-hole cross-sectional area in a single experiment, providing a comprehensive analysis of the effect of through-hole cross-sectional area on cell migration.

[0021] In embodiments, the substrate may comprise a set of through-holes organized such that, along at least a second direction parallel to the substrate, each successive through-hole is of equal or larger width relative to an immediately preceding through-hole if present, culminating in a final through-hole having a width greater than that of at least another through-hole in that direction. This embodiment allows for the study of cell migration across a range of through-hole widths in a single experiment, providing a comprehensive analysis of the effect of through-hole width on cell migration. In embodiments, the substrate may comprise a set of through-holes organized such that, along at least a second direction parallel to the substrate, each successive through-hole is of equal or larger hydraulic diameter relative to an immediately preceding through-hole if present, culminating in a final through-hole having a hydraulic diameter greater than that of at least another through-hole in that direction. This embodiment allows for the study of cell migration across a range of through-hole diameters in a single experiment, providing a comprehensive analysis of the effect of through-hole hydraulic diameter on cell migration.

[0022] In embodiments, the said set of through-holes may be organized such that, along said at least a second direction parallel to the substrate, each successive through-hole is separated from an immediately preceding through-hole, if present, by a distance which is either equal or larger relative to the distance separating the preceding through-hole from its own preceding through-hole, if present, and wherein the distance between the last through-hole and an immediately preceding through-hole is larger than that of at least another through-hole pair in that direction. This embodiment facilitates the differentiation of migration events based on through-hole geometry and spacing, enhancing the precision of the study.

[0023] In embodiments, the first and second sides of the second chamber may be parallel to each other and said at least a first direction parallel to the substrate is a direction perpendicular to the first and second sides of the second chamber. This embodiment ensures a good control over the gradient of the chemical substance in the second chamber.

[0024] In embodiments, said at least a first direction and said at least a second direction are perpendicular to each other. This maximizes the number of different combinations through-hole geometry / chemical substance concentration. This maximizes the number of different combinations through-hole geometry / chemical substance concentration.

[0025] Typically, said at least a first direction is a single first direction.

[0026] Typically, said at least a second direction is a single second direction.

[0027] Typically, said at least a first direction is a single first direction and said at least a second direction is a single second direction.

[0028] In embodiments, the delivery system may be a microfluidic system.

[0029] In embodiments, the microfluidic system may comprise a first microfluidic channel for delivering the chemical substance from the first side and a second microfluidic channel for delivering the chemical substance from the second side, each microfluidic channel comprising an inlet for fluids, an outlet for fluids, and a channel fluidly connecting the inlet and outlet, said channel comprising a plurality of openings in fluid communication with the second chamber, said openings being adapted for letting fluid exit the channel. This embodiment provides the advantage of precise control over the delivery of the chemical substance, enabling the establishment of complex gradient profiles. In embodiments, the channel comprising a plurality of openings can comprise a channel separated from the second chamber by a plurality of aligned protrusions, wherein the space between the protrusions are the openings. The protrusions may, in embodiments, connect a bottom and a top of the channel. In embodiments, the protrusions may be pillars. The openings are adapted for letting fluid exit the channel a cross to the second chamber. In embodiments, the width of the openings (e.g., the space between the protrusions) can be from 50 to 250 pm.

[0030] In embodiments, the openings may be adapted for letting fluid exit the channel while simultaneously preventing a biocompatible matrix (typically a gel, such as a hydrogel) typically present in the second chamber from entering the channel. This embodiment prevents the clogging of the microfluidic system and maintains the integrity of the chemical gradient.

[0031] In embodiments, the microfluidic system may further comprise driving means for continuously driving a fluid from the inlet to the outlet of the at least a first microfluidic channel. The driving means can be a syringe pump or any other type of pump.

[0032] In embodiments, the delivery system may be configured to establish a gradient of the chemical substance with a steepness adjustable by controlling the flow rates and / or concentration of the chemical substance through the first and second microfluidic channels. This embodiment allows for the dynamic adjustment of the chemical gradient, offering flexibility in experimental design.

[0033] In embodiments, the detection means may comprise a plurality of electrodes positioned adjacent to each through-hole for measuring electrical signal changes, such as impedance changes, associated with cell migration at that through-hole. In other words, the detection means may comprise a plurality of electrodes for measuring electrical signal changes associated with cell migration, said electrodes being positioned so that each through-hole has at least one electrode adjacent thereto. This embodiment enables the local, non-invasive, and real-time detection of cell migration events. It is an advantage of embodiments of the present invention that direct read-out of migration events is possible without the need for imaging-based quantifications, which can be labor- intensive and time-consuming.

[0034] In embodiments, the electrodes may have a width of from 1 to 50 pm. This embodiment ensures that the electrodes are appropriately sized for detecting the electrical signals associated with the cells of interest. In embodiments, the electrodes may be made of a biocompatible conductive material selected from the group consisting of gold, platinum, titanium nitride, and indium tin oxide. This embodiment ensures biocompatibility and reliable electrical signal detection.

[0035] In embodiments, at least part of the device may be optically transparent. This embodiment allows for the use of optical imaging techniques to complement the electrical detection of cell migration. In embodiments, the part of the device that is optically transparent is such as to allow for the optical detection of cell migration through the substrate. For instance, the top of the first chamber and / or the bottom of the second chamber may be transparent.

[0036] In embodiments, the detection means may include an optical imaging system configured to capture through said part of the device which is optically transparent images of cells migrating through the through-holes. This embodiment provides the advantage of visual confirmation and additional data on cell migration.

[0037] In embodiments, the optical imaging system may comprise a microscope with a camera. This embodiment provides high-resolution imaging capabilities for detailed analysis of cell migration.

[0038] In embodiments, the optical imaging system may be further adapted for performing time-lapse imaging to monitor cell migration over time. This embodiment allows for the study of cell migration dynamics and the effects of temporal changes in the chemical gradient.

[0039] In embodiments, the substrate may comprise a semiconductor material, e.g., silicon. Preferably, it is made of a rigid semiconductor material. More preferably, it is made of silicon. This embodiment provides the advantage of using a material with excellent mechanical and chemical stability, as well as compatibility with microfabrication techniques. It is an advantage of embodiments of the present invention that, when a semiconductor substrate is used, it provides high precision in the design and manufacture of through-holes down to the nanometer level, which is not typically as easy to achieve with polymer-based systems. In embodiments, the use of a silicon substrate enables the analysis of cell migration at the single-cell level, a level of precision not achievable with polymer-based devices. It is an advantage of embodiments of the present invention that the substrate can be made of silicon, which is known for its biocompatibility and mechanical stability, making it suitable for long-term cell culture experiments.

[0040] In embodiments, the through-holes may have shapes selected from the group consisting of circular, oval, rectangular, and irregular shapes. This embodiment allows for the study of the effect of through-hole geometry on cell migration. In embodiments, the first chamber may house biological cells comprising endothelial cells covering the substrate, and the second chamber may house target tissue cells and / or biocompatible matrix such as a gel, and preferably a hydrogel. This embodiment provides a model system for studying cell migration in a context that mimics physiological conditions.

[0041] In embodiments, the target tissue cells may be brain cells, e.g., neurons and the device may mimic the blood-brain barrier. This embodiment provides a specialized model for studying cell migration in the context of the blood-brain barrier.

[0042] In embodiments, the width or hydraulic diameter of the through-holes may be from 100 nm to 500 pm, preferably from 200 nm to 200 pm, more preferably from 300 nm to 100 pm, and most preferably from 1 pm to 30 pm. This embodiment allows for the study of cell migration through a wide range of through-hole width, relevant to various biological contexts. Cells should typically not be able to migrate through a through-hole of 100 nm. The smallest bacteria have a diameter of 200 nm, and eukaryotic cells are at least 800 nm large. Bacteria have limited ability to squeeze through holes smaller than themselves. Having holes as small as 100 nm has the advantage to test the translocation of the smallest bacteria when they are very attracted toward the other side of the substrate. In most case, however, a lower limit at 100 nm will not be useful and a lower limit at 200 nm, or even at 300 nm will be sufficient to study the translocation of bacteria. To study the transmigration of cells of the human or animal body, a lower limit at 1 pm is advantageous to test the transmigration of the smallest body cells when they are very attracted toward the other side of the substrate. Since the smallest body cells are about 6 pm in diameter, a through-hole having a width or hydraulic diameter of 1 pm would require the body cell to squeeze through the hole. The largest through-hole do not need to be larger than 500 pm since most cells of interest are not as large. Amongst cells of interest, some cancer cells can reach 500 pm.

[0043] In embodiments, the delivery system may include a reservoir for the chemical substance, the reservoir being in fluid communication with the first and second microfluidic channels. This embodiment provides a consistent and reliable source of the chemical substance for gradient establishment.

[0044] In embodiments, the device may further comprise a control system for regulating the delivery of the chemical substance to establish the desired uniform or gradient fields. This embodiment provides precise control over the experimental conditions, enhancing the reproducibility and accuracy of the study.

[0045] In embodiments, the device may further comprise a detection system configured to measure the concentration of the chemical substance at various locations of the second chamber, preferably at the level of the substrate. This embodiment allows for the verification and fine-tuning of the chemical gradient, ensuring that the experimental conditions are as intended.

[0046] In embodiments, the substrate may be removable without altering the rest of the device, thereby facilitating the replacement of the substrate to allow for multiple experimental runs with different through-hole configurations. This embodiment provides the advantage of reusability and adaptability for a range of experiments.

[0047] In embodiments, the substrate may be rigid, preferably entirely made of rigid materials, preferably entirely made of rigid silicon. This embodiment ensures structural integrity and durability of the device.

[0048] In embodiments, the device may comprise means for keeping constant the environmental conditions therein. Environmental conditions, such as temperature, pH, and humidity, are preferably maintained constant to ensure that observed effects on cell migration are due to the intended experimental parameters.

[0049] In the second aspect, the present invention relates to a system for studying cell migration, comprising the device of any embodiment of the first aspect comprising detection means and a data processing unit for analyzing data gathered with the detection means.

[0050] In embodiments, the data processing unit may be configured for quantifying the rate and volume (or amount) of cell migration through the through-holes. This embodiment provides the advantage of detailed quantitative analysis of cell migration, which is advantageous for understanding the underlying mechanisms.

[0051] In embodiments, the data processing unit may be configured to analyze output of the detection means, e.g., of the electrodes, to perform said quantification of the rate and / or volume (or amount). In embodiments, the data processing unit may be configured to analyze images captured by the detection means to perform said quantification of the rate and / or volume (or amount). In embodiments, the data processing unit may be configured to analyze images captured by the detection means to detect changes in cell morphology. These embodiments allow for the comprehensive analysis of cell behavior during migration.

[0052] In embodiments, the data processing unit may be configured to integrate data from the detection means and detection system. This embodiment provides a holistic view of the experimental data, enabling more accurate interpretations and conclusions.

[0053] In embodiments, the data processing unit may be further configured to perform statistical analysis to validate the presence of coupling or interdependence between parameters. This embodiment provides the advantage of rigorous data analysis, which is advantageous for scientific research.

[0054] In embodiments, the data processing unit may be configured to adapt a microfluidic flow or cell input once a change in gradient is detected in the second chamber. This can improve the reliability of the gradient conditions.

[0055] In the fourth aspect, the present invention relates to a process for manufacturing a device for studying cell migration, the process comprising providing a substrate, forming a set of through-holes in the substrate, wherein the through-holes comprise at least two types differing in their geometries, forming a first chamber on one side of the substrate and a second chamber on the opposite side, and integrating a delivery system capable of establishing both uniform and gradient fields of a chemical substance.

[0056] In embodiments, forming the set of through-holes may involve a photolithography process to create a pattern on the substrate followed by an etching process to generate the through-holes. This embodiment provides the advantage of precise and customizable through-hole creation, advantageous for the device's functionality.

[0057] In the fifth aspect, the present invention relates to a method for studying cell migration using the device of any embodiment of the first aspect or the system of any embodiment of the second aspect, the method comprising forming a cell culture into the first chamber, establishing a chemical substance gradient between the first and the second side of the second chamber using the delivery system, and detecting cell migration through the through-holes. In embodiments, the method further comprises providing a biocompatible matrix in the second chamber and providing target cells in the biocompatible matrix before to establish the chemical substance gradient.

[0058] It is an advantage of embodiments of the present invention that a comprehensive understanding of cell transmigration and pathogen translocation across various biological barriers can be facilitated. It is a further advantage of embodiments of the present invention that the influence of multiple parameters on cellular migration can be monitored simultaneously, providing insights into the complex interplay of chemical and physical cues that govern cell behavior.

[0059] It is an advantage of embodiments of the present invention that in situ evaluation of migration events, e.g., transmigration or translocation events, as a function of a chemical signal, whether static or dynamic, can be conducted.

[0060] It is an advantage of embodiments of the present invention that physiological and pathological fluid flow rates can be recreated, providing a more accurate simulation of in vivo conditions. It is an advantage of embodiments of the present invention that the device can mimic complex biological systems such as the blood-brain barrier, providing a platform for studying the transmigration of immune cells and the translocation of pathogens in a controlled environment. It is an advantage of embodiments of the present invention that the system can be used to explore the effects of chemotaxis on cell migration, with the ability to adjust the steepness of the chemical gradient through microfluidic control systems.

[0061] It is an advantage of embodiments of the present invention that the device can house both endothelial cells and target tissue cells, allowing for the study of interactions between different cell types within the same experimental setup.

[0062] Particular and preferred aspects of the invention are set out in the accompanying independent and dependent claims. Features from the dependent claims may be combined with features of the independent claims and with features of other dependent claims as appropriate and not merely as explicitly set out in the claims.

[0063] The above and other characteristics, features and advantages of the present invention will become apparent from the following detailed description, taken in conjunction with the accompanying drawings, which illustrate, by way of example, the principles of the invention. This description is given for the sake of example only, without limiting the scope of the invention. The reference figures quoted below refer to the attached drawings.

[0064] Brief description of the drawings

[0065] Fig.1 is a schematic representation of a device for studying cell migration, showing a range of through-hole width (D1 to D4) in ascending order according to embodiments of the present invention.

[0066] Fig.2 is a comparative illustration of static versus dynamic chemotaxis induced on a device for studying cell migration, demonstrating the difference in chemical gradient field generation according to embodiments of the present invention.

[0067] In the different figures, the same reference signs refer to the same or analogous elements.

[0068] Detailed description of Illustrative Embodiments

[0069] The present invention will be described with respect to particular embodiments and with reference to certain drawings, but the invention is not limited thereto but only by the claims. The drawings described are only schematic and are non-limiting. In the drawings, the size of some of the elements may be exaggerated and not drawn on scale for illustrative purposes. The dimensions and the relative dimensions do not correspond to actual reductions to practice of the invention.

[0070] The terms first, second, third and the like in the description and in the claims, are used for distinguishing between similar elements and not necessarily for describing a sequence, either temporally, spatially, in ranking or in any other manner. It is to be understood that the terms so used are interchangeable under appropriate circumstances and that the embodiments of the invention described herein are capable of operation in other sequences than described or illustrated herein.

[0071] Moreover, the terms top and over and the like in the description and the claims are used for descriptive purposes and not necessarily for describing relative positions. It is to be understood that the terms so used are interchangeable under appropriate circumstances and that the embodiments of the invention described herein are capable of operation in other orientations than described or illustrated herein.

[0072] It is to be noticed that the term “comprising”, also used in the claims, should not be interpreted as being restricted to the means listed thereafter; it does not exclude other elements or steps. It is thus to be interpreted as specifying the presence of the stated features, integers, steps or components as referred to, but does not preclude the presence or addition of one or more other features, integers, steps or components, or groups thereof. Thus, the scope of the expression “a device comprising means A and B” should not be interpreted as being limited to devices consisting only of components A and B. It means that with respect to the present invention, the only relevant components of the device are A and B. The term “comprising” therefore covers the situation where only the stated features are present and the situation where these features and one or more other features are present. The word “comprising” according to the invention therefore also includes as one embodiment that no further components are present. When the word "comprising" is used to describe an embodiment in this application, it is to be understood that an alternative version of the same embodiment, wherein the term "comprising" is replaced by "consisting of", is also encompassed within the scope of the present invention.

[0073] Similarly, it is to be noticed that the term “coupled” should not be interpreted as being restricted to direct connections only. The terms “coupled” and “connected”, along with their derivatives, may be used. It should be understood that these terms are not intended as synonyms for each other. Thus, the scope of the expression “a device A coupled to a device B” should not be limited to devices or systems wherein an output of device A is directly connected to an input of device B. It means that there exists a path between an output of A and an input of B which may be a path including other devices or means. “Coupled” may mean that two or more elements are either in direct physical or electrical contact, or that two or more elements are not in direct contact with each other but yet still co-operate or interact with each other.

[0074] Reference throughout this specification to “one embodiment” or “an embodiment” means that a particular feature, structure or characteristic described in connection with the embodiment is included in at least one embodiment of the present invention. Thus, appearances of the phrases “in one embodiment” or “in an embodiment” in various places throughout this specification are not necessarily all referring to the same embodiment, but may. Furthermore, the particular features, structures or characteristics may be combined in any suitable manner, as would be apparent to one of ordinary skill in the art from this disclosure, in one or more embodiments.

[0075] Similarly, it should be appreciated that in the description of exemplary embodiments of the invention, various features of the invention are sometimes grouped together in a single embodiment, figure, or description thereof for the purpose of streamlining the disclosure and aiding in the understanding of one or more of the various inventive aspects. This method of disclosure, however, is not to be interpreted as reflecting an intention that the claimed invention requires more features than are expressly recited in each claim. Rather, as the following claims reflect, inventive aspects lie in less than all features of a single foregoing disclosed embodiment. Thus, the claims following the detailed description are hereby expressly incorporated into this detailed description, with each claim standing on its own as a separate embodiment of this invention.

[0076] Furthermore, while some embodiments described herein include some, but not other features included in other embodiments, combinations of features of different embodiments are meant to be within the scope of the invention, and form different embodiments, as would be understood by those in the art. For example, in the following claims, any of the claimed embodiments can be used in any combination.

[0077] Furthermore, some of the embodiments are described herein as a method or combination of elements of a method that can be implemented by a processor of a computer system or by other means of carrying out the function. Thus, a processor with the necessary instructions for carrying out such a method or element of a method forms a means for carrying out the method or element of a method. Furthermore, an element described herein of an apparatus embodiment is an example of a means for carrying out the function performed by the element for the purpose of carrying out the invention.

[0078] In the description provided herein, numerous specific details are set forth. However, it is understood that embodiments of the invention may be practiced without these specific details. In other instances, well-known methods, structures and techniques have not been shown in detail in order not to obscure an understanding of this description.

[0079] The following terms are provided solely to aid in the understanding of the invention.

[0080] As used herein, and unless otherwise specified, the term "device for studying cell migration" refers to an apparatus specifically designed to enable the detection of the movement of cells from one location to another. In embodiments, the device can be for studying cell transmigration or cell translocation, wherein transmigration is a specific type of cell migration that involves the movement of cells of the human or animal body (e.g., immune cells or tumor cells) across a cell barrier, such as the endothelium, and wherein cell translocation, involves migration of pathogens or bacteria across a cell barrier, e.g. the intestinal barrier. This detection can be performed by any suitable detection means such as optical means and / or electrical means. In embodiments, optical means could comprise a microscope and / or transparent parts of the device enabling the observation (e.g., with the microscope) of the migration of cells through the substrate. Preferably, in addition to be adapted to detect the movement of the cells, the device is adapted for measuring the movement of cells through the substrate. This measurement can be a recording over time of the detections performed by the detection means. Means to perform this recording may, therefore, be present. In one aspect, the device is part of a system adapted to analyze the migration of the cells by analyzing data gathered by the detection means. The analysis can, for instance, be performed by a data processing unit. A device for studying cell migration can, therefore, be a device for detecting, measuring, and / or analyzing cell migration. This includes, but is not limited to, devices that can create chemical gradient fields to stimulate cell movement, contain chambers for cell culture, and have substrates with through-holes for cells to migrate through.

[0081] As used herein, and unless otherwise specified, the term "first chamber" refers to a designated compartment or space within the device that is intended to house or contain cells for the purpose of studying their migration. This chamber is part of the overall structure of the device and is positioned in such a way that it can interact with other components, such as a second chamber or a substrate with through-holes. As an example, in the case of a device mimicking the blood-brain barrier, said device being for studying cell migration through said barrier, the first chamber can be for the circulation of blood and the substrate may be lined with endothelial cells.

[0082] As used herein, and unless otherwise specified, the term "second chamber" refers to another designated compartment or space within the device that is separate from but related to the first chamber. It is designed to interact with the first chamber via a substrate that separates them and contains through-holes for fluidic connection and cell migration. The second chamber has a first side and a second side, which can be opposite to one another or have other spatial relationships depending on the design of the device. As an example, in the case of a device mimicking the blood-brain barrier, said device being for studying cell migration through said barrier, the second chamber can be for simulating the brain side of the blood-brain barrier and may comprise a biocompatible matrix (e.g., a hydrogel) and brain cells in the hydrogel.

[0083] As used herein, and unless otherwise specified, the term "substrate" refers to a material layer or structure that serves as a boundary between the first and second chambers. It includes a set of through-holes that allow for fluidic connection and the passage of cells between the chambers. The substrate can be made of various materials, including but not limited to silicon, and can have different properties such as being optically transparent or rigid.

[0084] As used herein, and unless otherwise specified, the term "set of through-holes" refers to multiple apertures or openings in the substrate that permit the flow of fluids and the migration of cells between the first and second chambers. This set includes at least two types of through-holes that differ in their geometries, i.e. , in their sizes (width, cross- sectional area, or hydraulic diameter) and / or shapes, which can be circular, oval, rectangular, or irregular, and can be organized in specific patterns to create desired migration paths or fluidic connections.

[0085] As used herein, unless specified otherwise, the term 'geometries' encompasses the shapes and physical dimensions of through-holes. Physical dimensions can, for instance, be selected from the list consisting of diameter, hydraulic diameter (i.e., four time the cross-sectional area divided by the perimeter), length, width, and cross-sectional area. The shape is the specific contours defining the perimeter. Preferably, the geometry of each through-hole may be determined at the top surface of the substrate, i.e., the side of the substrate facing the first chamber. Preferably, the geometry of each through-hole may be constant or substantially constant across the thickness of the substrate. In embodiments, each through-hole has a cross-section not varying by more than 10%, preferably not varying by more than 5%, around an average cross-section across the thickness of the substrate. The at least two types of through-holes comprised in the substrate may differ in physical dimensions, shape, or a combination of both. Example of shapes are circular, elliptical, rectangular, or any irregular form. These geometric distinctions are pivotal as they significantly influence the fluidic connectivity between the first chamber (100) and the second chamber (102), impacting cell migration behavior and the diffusion or transport of chemical substances (135) across the substrate. Leveraging variations in through-hole geometries enables the creation of targeted conditions for in- depth investigation of cell migration dynamics. As used herein, and unless otherwise specified, the term "delivery system" refers to a mechanism or assembly within the device that is capable of introducing and controlling the distribution of a chemical substance within the device. This system is capable to create both uniform and gradient fields of the chemical substance, which can influence cell migration. Preferably, a gradient field is created. The delivery system can include microfluidic components such as channels, inlets, outlets, and openings for precise fluid handling.

[0086] As used herein, and unless otherwise specified, the term "detection means" refers to any component or assembly within the device that is used to monitor, detect, or measure the migration of cells through the through-holes. This can include a variety of sensors or imaging systems, such as electrodes for measuring electrical signal changes or optical systems for capturing images of migrating cells.

[0087] As used herein, and unless otherwise specified, the term "microfluidic system" refers to a subset of the delivery system that utilizes microscale channels and other components to control the flow of fluids within the device. This system is designed to handle small volumes of fluids with precision and can include features such as programmable pumps and valves for adjusting chemical gradient fields dynamically.

[0088] As used herein, and unless otherwise specified, the term "biocompatible matrix" refers to a substance or material that is compatible with biological cells and can be used within the device to mimic the natural environment of tissues. Examples of biocompatible matrices include gels such as hydrogels and other materials that can support cell growth and migration.

[0089] As used herein, and unless otherwise specified, the term "chemical substance (135)" refers to any substance or compound that can be delivered by the delivery system to establish uniform or gradient fields within the second chamber. Examples of chemical substances include, but are not limited to, growth factors, cytokines, chemokines, and other signaling molecules.

[0090] As used herein, and unless otherwise specified, the term "control system" refers to a set of components within the device that regulates the operation of the delivery system. This system can adjust parameters such as flow rates (e.g., of the chemical substance) and chemical substance concentrations to establish the desired chemical fields within the device.

[0091] As used herein, and unless otherwise specified, the term "data processing unit" refers to hardware, or a combination of hardware and software that is used to analyze data gathered by the detection means. This unit can perform functions such as quantifying cell migration rates, analyzing images for migration events and changes in cell morphology, and performing statistical analysis to validate experimental results.

[0092] The invention will now be described by a detailed description of several embodiments of the invention. It is clear that other embodiments of the invention can be configured according to the knowledge of persons skilled in the art without departing from the technical teaching of the invention, the invention being limited only by the terms of the appended claims.

[0093] In the first aspect, the present invention relates to a device (1) for studying cell migration, comprising: a. A first chamber (100), b. A second chamber (102) having a first (104) and a second sides (106), c. A substrate (108) delimiting the first chamber (100) from the second chamber (102) and comprising a set of through-holes (110) for fluidically connecting the first chamber (100) and the second chamber (102), said set comprising at least two types of through-holes (110) differing in their geometries, and d. A delivery system (112) adapted for delivering a chemical substance (135) from the first (104) and the second sides (106), thereby enabling the establishment of both uniform fields and gradient fields of said chemical substance (135) between the first side (104) and the second side (106) in a first direction parallel to the substrate (108). The delivery system (112) is adapted for delivering a chemical substance (135) from the first (104) and the second sides (106), thereby enabling the establishment of a uniform field of said chemical substance (135) between the first side (104) and the second side (106) in a first direction parallel to the substrate (108). The delivery system (112) is adapted for delivering a chemical substance (135) from the first (104) and the second sides (106), thereby enabling the establishment of a gradient field of said chemical substance (135) between the first side (104) and the second side (106) in a first direction parallel to the substrate (108). The device permits the use to choose between establishing a uniform field or a gradient field. A gradient field enable the study of cell migration in a variety of chemical substance concentration at once.

[0094] The top of Fig. 1 shows a schematic cross-sectional side view of a device (1) for studying cell migration according to embodiments of the present invention. The device (1) comprises a substrate (108) with a set of through-holes (110) of varying geometries. In the top view of the substrate illustrated at the bottom of the figure, the through-holes (110) are organized such that, along a direction parallel to the substrate (108), each successive through-hole is of increasing width (or hydraulic diameter or cross-sectional area) relative to the immediately preceding through-hole, culminating in a final through-hole having the largest width (or hydraulic diameter or cross-sectional area). The through-holes (110) are labeled D1 to D4 to indicate their ascending order of dimensions. In embodiments, as illustrated in figures 1 and 2, the first (104) and second sides (106) may be opposite to one another. This configuration facilitates for the establishment of controlled chemical gradient fields across the substrate (108). In a further embodiment, two chemical gradients with different chemicals may be created in the second chamber (102). One gradient can be in an opposed, but parallel, direction to the other.

[0095] Fig. 2 shows a cross-sectional view illustrating static versus dynamic chemotaxis induced on the device (1) for exploring cell transmigration according to embodiments of the present invention. The device (1) comprises a first chamber (100) and a second chamber (102) separated by the substrate (108) with the set of through-holes (110). The second chamber (102) has a first side (104) and a second side (106) opposite to one another. As shown at the top of Fig. 1 , a delivery system (112) is integrated into the device (1) for delivering a chemical substance (135) from the first side (104) and the second side (106), thereby enabling the establishment of both uniform fields and gradient fields of the chemical substance (135) between the first side (104) and the second side (106) in a first direction parallel to the substrate (108). In the static chemotaxis setup (bottom), the chemical substance (135) is uniformly distributed in the second chamber (102). In the dynamic chemotaxis setup (top), the delivery system (112) establishes a gradient of the chemical substance (135) in the second chamber (102).

[0096] In embodiments, the device may further comprise detection means (114) for detecting the migration of cells through each through-hole (110). This enables quantitative analysis of cell migration behavior. The detection means (114) may comprise a plurality of electrodes (128) positioned adjacent to each through-hole (110) for measuring electrical signal changes, such as impedance changes, associated with cell migration at that through-hole (110). Electrical sensing enables label-free detection of migration events. The electrodes (128) may have dimensions ranging from 1-50 pm. This size range is suitable for detecting individual cells. The electrodes (128) may be made of a biocompatible conductive material selected from the group consisting of gold, platinum, titanium nitride, and indium tin oxide. These materials are compatible with cell culture and provide good electrical conductivity.

[0097] In embodiments, at least part of the device may be optically transparent. Optical transparency allows for visual inspection and imaging of the cells. The detection means (114) may include an optical imaging system configured to capture through said part of the device which is optically transparent images of cells migrating through the through- holes (110). Imaging provides detailed information about cell morphology and behavior. The optical imaging system may comprise a microscope with a camera. This setup enables high-resolution imaging. The optical imaging system may be further adapted for performing time-lapse imaging to monitor cell migration over time. Time-lapse imaging reveals the dynamics of the migration process.

[0098] In embodiments, the substrate (108) may comprise a set of through-holes (110) organized such that, along at least a second direction (103) parallel to the substrate (108), each successive through-hole is of equal or increasing width (or hydraulic diameter or cross-sectional area) relative to an immediately preceding through-hole if present, culminating in a final through-hole having a width (or hydraulic diameter or cross-sectional area) greater than that of at least another through-hole in that direction. This gradient in through-hole size allows for the study of size-dependent migration.

[0099] In embodiments, the said set of through-holes (110) may be organized such that, along said at least a second direction (103) parallel to the substrate (108), each successive through-hole is separated from an immediately preceding through-hole, if present, by a distance which is either equal or increasing relative to the distance separating the preceding through-hole from its own preceding through-hole, if present, and wherein the distance between the last through-hole and an immediately preceding through-hole is larger than that of at least another through-hole pair in that direction. This spatial gradient provides additional control over migration behavior.

[0100] In embodiments, the first (104) and second sides (106) of the second chamber (102) may be parallel to each other and said at least a first direction parallel to the substrate (108) may comprise a direction perpendicular to the first (104) and second sides (106) of the second chamber (102). This allows for the establishment of linear chemical gradient fields.

[0101] In embodiments, the delivery system (112) may be a microfluidic system. Microfluidics enable precise control over the chemical environment. The microfluidic system may comprise a first microfluidic channel (116) for delivering the chemical substance (135) from the first side (104) and a second microfluidic channel (118) for delivering the chemical substance (135) from the second side (106), each microfluidic channel comprising an inlet (120) for fluids, an outlet (122) for fluids, and a channel (124) fluidly connecting the inlet (120) and outlet (122), said channel (124) comprising a plurality of openings (126) in fluid communication with the second chamber (102), said openings (126) being adapted for letting fluid exit the channel. This configuration allows for the controlled release of chemicals into the second chamber (102). The delivery system (112) may be configured to establish a gradient of the chemical substance (135) with a steepness adjustable by controlling the flow rates and / or concentration of the chemical substance (135) through the first (116) and second microfluidic channels (118). This provides flexibility in gradient generation. The openings (126) may be adapted for letting fluid exit the channel while simultaneously preventing a biocompatible matrix (e.g., a hydrogel) from entering the channel. This maintains the integrity of the biocompatible matrix in the second chamber (102), when present. The delivery system (112) may include a reservoir for the chemical substance (135), the reservoir being in fluid communication with the first (116) and second microfluidic channels (118). The reservoir provides a source of the chemical for gradient generation.

[0102] In embodiments, the substrate (108) may comprise silicon, preferably is made of silicon. Silicon is a robust and precise material for microfabrication. The through-holes (110) may have any shape and may, for instance, be selected from the group consisting of circular, oval, rectangular, and irregular shapes. Different shapes can influence migration behavior. The width (or hydraulic diameter or cross-sectional area) of the through-holes (110) may be from 100 nm to 500 pm, preferably 1 pm to 200 pm. This size range covers the most relevant dimensions for cell migration.

[0103] In embodiments, the first chamber (100) may house biological cells comprising endothelial cells (105) covering the substrate (108), and the second chamber (102) may house target tissue cells (107) and / or biocompatible matrix such as a hydrogel. This setup mimics the physiological environment of cell migration. The target tissue cells may be neurons and the device may mimic the blood-brain barrier. This allows for the study of neuronal migration in a relevant context.

[0104] In embodiments, the device may further comprise a control system for regulating the delivery of the chemical substance (135) to establish the desired uniform or gradient fields. Automated control enables dynamic modulation of the chemical environment. The device may further comprise a detection system (133) configured to measure the concentration of the chemical substance (135) at various locations of the second chamber (102). This allows for real-time monitoring of the chemical gradient fields.

[0105] In embodiments, the substrate (108) may be removable without altering the rest of the device, thereby facilitating the replacement of the substrate (108) to allow for multiple experimental runs with different through-hole configurations. Removable substrates provide flexibility in experimental design. The substrate (108) may be rigid, preferably entirely made of rigid materials, preferably entirely made of rigid silicon. Rigidity ensures the stability of the through-hole structures.

[0106] Any element of the first aspect may be as correspondingly described in any embodiment of the other aspects.

[0107] In the second aspect, the present invention relates to a system for studying cell migration, comprising: i. the device of any one of the embodiments of the first aspect; ii. a data processing unit (134) for analyzing data gathered with the detection means (114). In embodiments, the data processing unit (134) may be configured for quantifying the rate and volume (or amount) of cell migration through the through-holes (110). This enables quantitative analysis of migration behavior. In embodiments, the data processing unit (134) may be configured for quantifying the rate and volume (or amount) of cell migration through the through-holes (110) within a predetermined time period. This allows for the analysis of migration dynamics over specific time scales.

[0108] The data processing unit (134) may be configured to analyze images captured by the detection means (114) to identify and quantify migration events and changes in cell morphology. Image analysis extracts valuable information from the visual data.

[0109] In embodiments, the delivery system (112) may include microfluidic control systems with programmable pumps and valves for dynamically adjusting the chemical gradient fields, e.g., in function of a detection system (133). Feedback control allows for adaptive modulation of the chemical environment. The data processing unit (134) may be configured to integrate data from the detection means (114) and detection system (133). Data integration provides a comprehensive view of the migration process. The data processing unit (134) may be further configured to perform statistical analysis to validate the presence of coupling or interdependence between parameters. Statistical analysis reveals the relationships between different experimental variables. For instance, the data processing unit (134) may be further configured to perform statistical analysis to validate the presence of coupling or interdependence between pore geometries and chemical substance concentration.

[0110] Any embodiment of the second aspect may be as correspondingly described in the first aspect.

[0111] In the third aspect, the present invention relates to a process for manufacturing a device for studying cell migration, the process comprising: a. Providing a substrate (108); b. Forming a set of through-holes (110) in the substrate (108), wherein the through-holes (110) comprise at least two types differing in their geometries; c. Forming a first chamber (100) on one side of the substrate (108) and a second chamber (102) on the opposite side; and d. Integrating a delivery system (112) capable of establishing both uniform and gradient fields of a chemical substance (135).

[0112] In embodiments, forming the set of through-holes (110) may involve a photolithography process to create a pattern on the substrate (108) followed by an etching process to generate the through-holes (110). Photolithography and etching are standard microfabrication techniques for creating precise structures.

[0113] Any embodiment of the third aspect may be as correspondingly described in the first or the second aspect. In the fourth aspect, the present invention relates to a method for studying cell migration using the device of any one of the embodiments of the first aspect or the system of the second aspect, the method comprising: a. Forming a cell culture into the first chamber (100); b. Establishing a chemical substance (135) gradient between the first (104) and the second side (106) of the second chamber (102) using the delivery system (112); c. Detecting cell migration through the through-holes (110).

[0114] The advantage of this method is that it allows for the comprehensive study of cell migration under controlled chemical gradient fields and through-holes of varying geometries, thereby providing insight into the complex interplay of factors that influence migration behavior in physiologically relevant contexts.

[0115] Any embodiment of the fourth aspect may be as correspondingly described in the other aspects.

[0116] Example 1 : A Multifunctional Silicon Chip for Studying Cell Transmigration and Translocation

[0117] A silicon chip is designed and fabricated to study cell transmigration and translocation as a function of through-hole size and chemical signaling. The chip consists of an upper chamber and a lower chamber separated by a porous silicon substrate. The through-holes in the substrate vary in size from 100 nm to hundreds of microns, distributed in a gradient pattern from the smallest to the largest through-holes. This gradient design allows for the simultaneous exploration of cell transmigration behavior across different through-hole sizes within a single chip.

[0118] To model the blood-brain barrier (BBB), endothelial cells are cultured on the upper side of the porous substrate to form a monolayer. Immune cells are then perfused through the first chamber using a microfluidic system, allowing them to interact with and potentially transmigrate through the endothelial layer. The second chamber of the chip is provided with a collagen hydrogel and populated with astrocytes or glial cells to represent the brain side of the BBB.

[0119] In addition to the through-hole size gradient, the chip is designed to investigate cell transmigration in response to chemical signaling or chemotaxis. Various molecules of interest (e.g., cytokines or chemokines), with different sizes and attractive properties, are introduced into the second chamber at desired concentrations. Two approaches are employed to establish chemical gradient fields: a static system and a dynamic system. In the static system, a fluidic connection is used to provide the chemical substance to the second chamber simultaneously from both sides thereof, until a homogeneous concentration is achieved throughout the second chamber, while in the dynamic system, a fluidic connection is used to provide one or multiple sources and sinks of the chemical substance, resulting in distinct concentration gradients.

[0120] Cell transmigration events are measured using two methods. First, the chip is optically transparent on both, the top side and the bottom side, allowing for imaging of cell migration from either the top or bottom of the device. Second, TiN electrodes are positioned around the through-holes in the middle plane of the chip. These electrodes, ranging in size from 1 to 20 pm and in various configurations (round, square, etc.), are used to detect transmigration events based on changes in electrical properties.

[0121] To enable continuous monitoring of the influence of multiple parameters simultaneously and to determine potential coupling or interdependence between these parameters, several additional features are integrated into the device. High-resolution time-lapse microscopy and live-cell imaging systems are employed to visualize cell migration in real-time, capturing dynamic changes in cell behavior in response to varying gradients and through-hole sizes. Automated image analysis software is used to process the vast amounts of imaging data, identifying and quantifying migration events, cell velocities, and morphological changes.

[0122] Precise control over chemical gradient fields is achieved through an integrated microfluidic system with programmable pumps and valves, allowing for the fine-tuning of chemical concentrations and the generation of complex gradient profiles. Environmental conditions, such as temperature, pH, and humidity, are maintained constant to ensure that observed effects on cell migration are due to the intended experimental parameters.

[0123] A comprehensive data management system is developed to integrate data from imaging, sensors, and microfluidic controls. Advanced statistical and computational models, including machine learning algorithms, are applied to analyze the interdependence and coupling of parameters, identifying complex patterns and interactions.

[0124] The chip design also incorporates a high-throughput capability, enabling parallel experiments under varying conditions. An array of microenvironments, each with its own set of conditions, is fabricated within a single device to allow for statistical validation of parameter coupling and interdependence.

[0125] The results obtained from this device provide valuable insights into cell transmigration and translocation behavior. The through-hole size gradient reveals a clear relationship between through-hole dimensions and the likelihood of cells successfully crossing the barrier. Smaller through-holes present a greater physical challenge for cells, resulting in lower transmigration rates compared to larger through-holes. The chemical signaling experiments demonstrate the influence of different molecules and concentration gradients on cell migration, as a function of through-hole size. The static and dynamic systems produce distinct gradient profiles, leading to variations in cell response and transmigration patterns. When a chemical substance gradient field is established in a first direction perpendicular to the second direction (103) of the through-hole size gradient, a maximal number of combination concentration / size is obtained. When a chemical substance gradient field is established in a first direction parallel to the second direction (103) of the through-hole size gradient, a smaller number of combination concentration / size is obtained but a plurality of observations in a same configuration is facilitated, thereby permitting a better understanding of the reproducibility of the observations.

[0126] The combination of real-time imaging and electrode-based detection allows for a comprehensive analysis of cell migration events. The imaging data provides detailed information on cell morphology, velocity, and interactions with the endothelial layer and through-holes. The electrode measurements complement the imaging results, enabling the quantification of transmigration events and the identification of any preferential migration paths.

[0127] The integration of multiple parameters within a single chip highlights the complex interplay between physical and chemical cues in regulating cell transmigration. The data management system and computational models reveal potential coupling and interdependence between through-hole size, chemical gradient fields, and cell behavior. These findings contribute to a deeper understanding of the factors governing cell migration in physiological and pathological processes.

[0128] Example 2: Evaluation of Cell Barrier Models on Transmigration Rates

[0129] The purpose of this experiment is to evaluate how different cell barrier models, such as endothelial or intestinal barriers, affect the rate of cell transmigration through the silicon chip. This will help in understanding the role of various biological barriers in physiological and pathological processes.

[0130] The silicon chip as described in example 1 is prepared with a range of through- hole shapes instead of different sizes. Different cell barriers are cultured on the chip to mimic various biological scenarios. A chemical gradient is established using the microfluidic system, and cell transmigration is induced. The migration of cells through the through-holes is detected using the integrated electrodes and optical imaging system. Data is collected and analyzed to determine the influence of the cell barrier model on transmigration rates. It is to be understood that although preferred embodiments, specific constructions and configurations, as well as materials, have been discussed herein for devices according to the present invention, various changes or modifications in form and detail may be made without departing from the scope of this invention. For example, any formulas given above are merely representative of procedures that may be used. Functionality may be added or deleted from the block diagrams and operations may be interchanged among functional blocks. Steps may be added or deleted to methods described within the scope of the present invention.

Claims

Claims1. A device for studying cell migration, comprising: a. A first chamber (100), b. A second chamber (102) having a first (104) and a second sides (106), c. A substrate (108) delimiting the first chamber (100) from the second chamber (102) and comprising a set of through-holes (110) for fluidically connecting the first chamber (100) and the second chamber (102), said set comprising at least two types of through-holes differing in their geometries, and d. A delivery system (112) adapted for delivering a chemical substance (135) from the first (104) and the second sides (106), thereby enabling the establishment of both uniform fields and gradient fields of said chemical substance (135) between the first side (104) and the second side (106) in a direction parallel to the substrate (108).

2. The device according to claim 1 , wherein the first (104) and second sides (106) are opposite to one another.

3. The device according to claim 1 or claim 2, further comprising detection means (114) for detecting the migration of cells through each through-hole.

4. The device of any one of the preceding claims, wherein the substrate (108) comprises a set of through-holes (110) organized such that, along at least a second direction (103) parallel to the substrate, each successive through-hole is of equal or increasing width, hydraulic diameter, or cross-sectional area relative to an immediately preceding through-hole if present, culminating in a final through-hole having a width, hydraulic diameter, or cross-sectional area, greater than that of at least another through- hole in that direction.

5. The device of claim 4, wherein the said set of through-holes (110) are organized such that, along said at least a second direction (103) parallel to the substrate, each successive through-hole is separated from an immediately preceding through-hole, if present, by a distance which is either equal or increasing relative to the distance separating the preceding through-hole from its own preceding through-hole, if present,and wherein the distance between the last through-hole and an immediately preceding through-hole is larger than that of at least another through-hole pair in that direction.

6. The device of claim 4 or claim 5, as depending on claim 2, wherein the first (104) and second sides (106) of the second chamber (102) are parallel to each other and said at least a first direction parallel to the substrate comprise a direction perpendicular to the first (104) and second sides (106) of the second chamber (102).

7. The device of any preceding claim, wherein the delivery system (112) is a microfluidic system, preferably comprising a first microfluidic channel (116) for delivering the chemical substance (135) from the first side (104) and a second microfluidic channel (118) for delivering the chemical substance (135) from the second side (106), each microfluidic channel comprising an inlet (120) for fluids, an outlet (122) for fluids, and a channel (124) fluidly connecting the inlet (120) and outlet (122), said channel (124) comprising a plurality of openings (126) in fluid communication with the second chamber (102), said openings (126) being adapted for letting fluid exit the channel.

8. The device of any of claims 3 to 7, wherein the detection means (114) comprises a plurality of electrodes (128) for measuring electrical signal changes associated with cell migration, said electrodes (128) being positioned so that each through-hole (110) has at least one electrode adjacent thereto.

9. The device of any preceding claim, wherein at least part of the device is optically transparent.

10. The device of any preceding claim, wherein the substrate (108) comprises silicon, preferably is made of silicon.

11. The device of any preceding claim, for studying cell transmigration.

12. The device of claim 11 , wherein the first chamber (100) houses biological cells comprising endothelial cells covering the substrate (108), and the second chamber (102) houses target tissue cells and / or biocompatible matrix such as a hydrogel.

13. The device of claim 12, wherein the target tissue cells are brain cells.

14. The device of any of the preceding claims, further comprising a detection system (133) configured to measure the concentration of the chemical substance (135) at various locations of the second chamber (102).

15. A system for studying cell migration, comprising: i. the device of any one of claims 3 to 14; and ii. a data processing unit (134) for analyzing data gathered with the detection means (114).

16. A method for studying cell migration using the device of any one of claims 1 to 13 or the system of claim 14, the method comprising: a. Forming a cell culture into the first chamber (100); b. Establishing a chemical substance (135) gradient between the first (104) and the second side (106) of the second chamber using the delivery system (112); and c. Detecting cell migration through the through-holes (110).

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