Digital microfluidics system

Abstract

The present disclosure relates to a digital microfluidics, 'DMF', system and a method for operating the same. The DMF system comprises: a first substrate unit comprising a first substrate having arranged thereon a plurality of electrodes for manipulation of a liquid volume; and a second substrate unit spaced apart from the first substrate unit in a first direction, wherein the second substrate unit comprises a second substrate having arranged thereon an organic electrochemical transistor, 'OECT', biosensor.

Description

[0001] DIGITAL MICROFLUIDICS SYSTEM

[0002] FIELD

[0003] The present disclosure generally relates to a digital microfluidics system and method of operating the same. More in particular, the present disclosure is especially suitable for ion concentration sensing in digital microfluidics for cell microenvironment monitoring.

[0004] BACKGROUND

[0005] Digital microfluidics (DMF) is a technique that enables automating biological experiments by precisely manipulating small liquid volumes, primarily using a concept known as electrowetting-on-dielectric (EWOD). Recently, DMF is becoming popular for customized immunological assays, cell-based assays, and DNA amplification. Various detection platforms are integrated for analytical purposes, such as optical, electrical, and mass spectrometry.

[0006] However, the bulkiness of conventional sensors, their limited utility outside of labs, and high fabrication costs impede wider application, particularly in point-of-care (POC) diagnostics. Despite the promise of electrical-based detection, issues remain in reproducibility and specificity.

[0007] Accordingly, there is a need for precise, reliable sensing devices to enhance DMF functionality in diagnostics.

[0008] SUMMARY

[0009] It is an object of the present disclosure to provide a DMF system in which the abovementioned problem(s) do not occur, or hardly so.

[0010] According to an aspect of the present disclosure, a digital microfluidics, ‘DMF’, system is provided, comprising: a first substrate unit comprising a first substrate having arranged thereon a plurality of electrodes for manipulation of a liquid volume (e.g., a (micro)droplet); and a second substrate unit spaced apart from the first substrate unit in a first direction, wherein the second substrate unit comprises a second substrate having arranged thereon an organic electrochemical transistor, ‘OECT’, biosensor.

[0011] The DMF system in accordance with the present disclosure addresses the challenges of precise and selective ion sensing in microfluidic environments by integrating OECT biosensors with DMF. Traditional methods, such as manual pipetting, are labour-intensive, time-consuming, and prone to errors. By combining the high sensitivity of OECTs with the automated droplet manipulation capabilities of DMF, the invention provides a scalable, versatile solution for real-timeion analysis, particularly in applications like cell culture monitoring. It offers a detection range from nanomolar to millimolar concentrations, enabling precise measurements across a wide spectrum of biological and chemical applications.

[0012] In an example, when viewed in the first direction, a position of a transistor of the OECT biosensor may overlap with at least one of the plurality of electrodes.

[0013] The second substrate may comprise a first portion and a second portion. The first portion and the second portion may each overlap with at least some of the plurality of electrodes. The first portion may include conductive material (e.g., a conductive layer).

[0014] The use of a conductive material in or on the second substrate, opposite the plurality of electrodes, can facilitate control and in particular movement of the liquid volume during operation.

[0015] The OECT biosensor may be arranged on the second portion. That is, the OECT biosensor may be integrated into the second substrate in such a manner that it does not inhibit the advantages of using conductive material as described above, or at least hardly so. For example, a majority of the plurality of electrodes may overlap with the first portion, and only a small subset of the plurality of electrodes (e.g., one or a few rows / columns when arranged in a matrix) may overlap with the second portion to allow sensing using the OECT biosensor, where the conductive material is not present to not inhibit the functionality of the OECT biosensor.

[0016] The first portion may be coated with the conductive material. Additionally or alternatively, the conductive material may comprise indium tin oxide, ‘ITO’. Additionally or alternatively, the second substrate may be made of glass. The first and second substrate may be made of the same or similar material, though this need not be the case. Additionally or alternatively, the conductive material may be configured to be electrically grounded during operation. That is, during operation, a positive voltage may be applied to a given electrode among the plurality of electrodes of the first substrate, whereas the conductive material, which may itself be considered an electrode of the second substrate, is electrically connected to a reference voltage, such as ground.

[0017] The first substrate may be coated with a first hydrophobic layer facing the second substrate. Additionally or alternatively, the second substrate may be coated with a second hydrophobic layer facing the first substrate. Such layers may enhance mobility of the liquid volume during DMF manipulation.

[0018] The first substrate unit may further comprise a dielectric layer between the first hydrophobic layer and the first substrate. For example, the DMF system may be configured to manipulate the liquid volume using electrowetting-on-dielectric, ‘EWOD’. However, the present disclosure is not limited to EWOD, and the dielectric layer as well as the hydrophobic layers may be omitted entirely, for example depending on the material of the first and second substrate.

[0019] The plurality of electrodes may be integrated on the first substrate. Additionally or alternatively, the transistor of the OECT biosensor may be integrated on the second substrate. Forexample, the transistor of the OECT biosensor and / or the plurality of electrodes may be fabricated on the second substrate or first substrate, respectively, through a (photo)lithography process.

[0020] The transistor of the OECT biosensor may comprise a source terminal, a drain terminal, and a gate terminal that are arranged in-plane. This compact configuration enables a reduction in size of the DMF system as a whole and thus may increase its practicability in, for example, small portable devices.

[0021] The OECT biosensor may be an ion-selective OECT, ‘IS-OECT’, biosensor including an ion-selective membrane. For example, the ion-selective membrane may be disposed over a channel of the transistor that extends between the source terminal and the drain terminal of the transistor. Accordingly, when a given gate voltage is applied to gate terminal of the transistor of the OECT biosensor, the ion-selective membrane may be permeable to (and thus allow passage of) a given ion or ions from the liquid volume to the channel of the transistor, while being impermeable to other ions which are therefore blocked by the ion-selective membrane. As a result, the conductivity of the channel may be a function of the ion concentration of the given ion(s) to which the ion-selective membrane is permeable. In this manner, the OECT biosensor is able to sense ion concentrations of specific ions in the liquid volume, depending on the ion-selective membrane used.

[0022] The DMF system may comprise plurality of said OECT biosensors. These may be identical or similar to each other in structure and arrangement and may enable the DMF system to measure, concurrently or sequentially, the same or different characteristics (e.g., ion concentrations of different ions) of the liquid volume.

[0023] For example, the plurality of IS-OECT biosensors may comprise a first IS-OECT biosensor with a first ion-selective membrane, and a second IS-OECT biosensor with a second ion-selective membrane. The first ion-selective membrane and the second ion-selective membrane may have different ion selectivity.

[0024] The plurality of electrodes may be arranged in a matrix of rows and columns. For example, the liquid volume may be configured to be manipulated to move in orthogonal directions. This can simplify the control of the plurality of electrodes for the movement of the liquid volume.

[0025] The first substrate unit may further comprises at least one reservoir electrode configured to hold an amount of liquid from which the liquid volume to be manipulated can be drawn by the plurality of electrodes. In a further example, a size of the at least one reservoir electrode may be greater than a size of each of the plurality of electrodes.

[0026] The DMF system may further comprise a control unit configured to actuate the plurality of electrodes in a given manner to move the liquid volume to a desired location between the first and second substrate. For example, the control unit may be integrally formed with the first substrate, or may be separate and connectable to the plurality of electrodes.The DMF system may further comprise a sensing unit configured to apply a gate voltage to the OECT biosensor and sense a corresponding current through the OECT biosensor. For example, the sensing unit may be integrally formed with the second substrate, or may be separate and connectable to the OECT biosensor.

[0027] The DMF system may further comprise a result deriving unit configured to determine a characteristic of the liquid volume based on the current through the OECT biosensor received from the sensing unit. For example, the characteristic may be an ion concentration present in the liquid volume, preferably of a given ion, such as Sodium, Potassium, and / or a pH level (i.e., an H+ and / or OH- concentration).

[0028] The first substrate unit and the second substrate unit may be formed as plates that extend in parallel to each other.

[0029] The DMF system may further comprise at least one further sensor at least partially integrated on the second substrate. For example, the at least one further sensor may include: one or more electrochemical sensors, such as an electrochemical impedance spectroscopy (EIS) based sensor, a field-effect transistor (FET) biosensor, or an aptamer-based biosensor potentiostat.

[0030] Additionally or alternatively, the at least one further sensor may include one or more optical sensors, such as a fiber-tip photonic crystals biosensor, a surface enhanced Raman scattering (SERS) sensor, or a surface plasmon resonance (SPR) sensor.

[0031] According to another aspect of the present disclosure, a wearable electronic device is provided, comprising the DMF system according to any of the embodiments above. Such a wearable device may enable automated or autonomous operation of the DMF system, for example for the purpose of real-time health monitoring.

[0032] For example, the DMF system may be part of a perspiration (i.e., sweat) sensor that can be worn on the skin of a user (e.g., a wrist), and may measure an ion concentration in liquid volume derived from sweat in an automated or autonomous manner.

[0033] According to another aspect of the present disclosure, an assembly is provided, comprising: a first substrate unit comprising a first substrate having arranged thereon a plurality of electrodes for digital microfluidics, ‘DMF’, based manipulation of a liquid volume; and a second substrate unit comprising a second substrate having arranged thereon an organic electrochemical transistor, ‘OECT’, biosensor.

[0034] According to yet another aspect of the present disclosure, a method for operating the DMF system described above is provided, the method comprising: a) providing the liquid volume between the first and second substrate unit; b) moving, using the plurality of electrodes, the liquid volume from a first position that is away from the OECT biosensor to a second position adjacent to the OECT biosensor in the first direction; and c) performing a measurement using the OECT biosensor.Step a) may further comprise providing cells in a medium between the first and second substrate at a third position different from the first and second position. Furthermore, step b) may comprise: moving, using the plurality of electrodes, the liquid volume from the first position to the third position; allowing the cells in the medium to exchange ions with the liquid volume; and moving, using the plurality of electrodes, the liquid volume from the third position to the second position.

[0035] Step a) may comprise: providing an amount of liquid on a reservoir electrode of the first substrate unit, the amount of liquid including the liquid volume; and acquiring, using the plurality of electrodes, the liquid volume from the amount of liquid to provide the liquid volume at the first position.

[0036] Further aspects and / or embodiments of the present disclosure may become apparent to those skilled in the art from the detailed description below.

[0037] BRIEF DESCRIPTION OF THE DRAWINGS

[0038] Next, the present disclosure will be described in more detail with reference to the appended drawings, wherein:

[0039] FIG. 1 is a simplified perspective view of a DMF system in accordance with an embodiment of the present disclosure;

[0040] FIG. 2A and 2B are top views of a first and second substrate of the DMF system, respectively, in accordance with the present disclosure;

[0041] FIG. 3 is a cross-sectional view of the DMF system in accordance with an embodiment of the present disclosure;

[0042] FIG. 4A-4L are cross-sectional diagrams illustrating a fabrication process for an OECT in accordance with the present disclosure;

[0043] FIG. 5 is a cross-sectional view of an OECT according to an embodiment of the present disclosure;

[0044] FIG. 6 is a flowchart illustrating a method for operating the DMF system, in accordance with an embodiment of the present disclosure;

[0045] FIG. 7A and 7B are diagrams illustrating measurement results for a Sodium-selective OECT according to an embodiment of the present disclosure; and

[0046] FIG. 8A and 8B are diagrams illustrating measurements results for a Potassium-selective OECT according to an embodiment of the present disclosure.

[0047] DETAILED DESCRIPTIONHereinafter, reference will be made to the appended drawings. It should be noted that identical reference signs may be used in different drawings to refer to identical or similar components. Moreover, unless explicitly stated otherwise, various elements shown in the appended drawings may not be drawn to scale. Rather, certain parts may be exaggerated or omitted for convenience of explanation.

[0048] Although repeated reference may be made to OECT-based ion sensing in particular, the present disclosure is not to be construed as being limited thereto. Functionalized OECT biosensors can additionally or alternatively be used for glucose, lactate, liposome, dopamine, DNA and bacteria detection as well as for ultrasensitive detection of proteins.

[0049] FIG. 1 illustrates a simplified view of a DMF system 100 according to an embodiment of the present disclosure. DMF system 100 includes first substrate unit 10 and second substrate unit 20, which are both formed as plates in this example. DMF system 100 may further include a control unit 40, a sensing unit 50, and / or a result deriving unit 60. For convenience of explanation, second substrate unit 20 is transparently drawn to enable viewing an underlying portion of first substrate unit 10.

[0050] First substrate unit 10 includes a first substrate 11 having a plurality of electrodes 14 arranged (e.g., integrated) thereon for liquid volume manipulation (i.e., movement). For example, electrodes 14 are arranged structurally in an array of rows and columns, enabling the liquid volume to be moved orthogonally.

[0051] First substrate 11 may also have at least one reservoir electrode 15 (eight shown in FIG. 1) arranged (e.g., integrated) thereon, adjacent to at least one of electrodes 14. During operation, an amount of liquid may be arranged on said reservoir electrode 15 from which the liquid volume to be manipulated can be withdrawn by electrodes 14.

[0052] First substrate 11 may include a ground electrode 16 which during operation is configured to be electrically grounded.

[0053] First substrate 11 may include a plurality of contact pads 17, each being connected to a respective electrode 14, reservoir electrode 15, or ground electrode 16, for example by wiring integrated on first substrate 11 or provided in a metal layer of first substrate unit 10. Only part of the wiring is illustrated in FIG. 1. Alternative implementations are also envisaged; for example, the wiring and / or contact pads 17 may at least partially be provided on a backside of first substrate 11 and may be electrically coupled to electrodes 14, reservoir using corresponding vias extending through first substrate 11.

[0054] Control unit 40 (or a controller external to DMF system 100) may be configured to actuate electrodes 14, reservoir electrodes 15, and ground electrode 16 by applying a given voltage to corresponding contact pads 17. In this manner, control unit 40 can control the operation of DMF system 100. For example, electrodes 14 may be actuated in a given manner to move the liquidvolume from one position between first substrate unit 10 and second substrate unit 20 to another by consecutively applying a voltage to adjacent electrodes 14.

[0055] Second substrate unit 20 includes a second substrate 21 having an OECT biosensor 30 arranged (e.g., integrated) thereon. In this example, second substrate 21 divided into a first portion 21a and a second portion 21b, and OECT biosensor 30 is arranged in particular on or at second portion 21b. For example, first portion 21a may be made of a conductive material (not shown in FIG. 1) and second portion 21b may be made of glass. Alternatively, both first portion 21 and second portion 21b are made of glass, but a conductive layer (not shown in FIG. 1) is provided on first portion 21a.

[0056] As shown in FIG. 1, a majority of electrodes 14 may be arranged to overlap first portion 21a, and only a small portion of electrodes 14 (e.g., only three rows in the example shown in FIG.

[0057] 1) may be arranged to overlap second portion 21b. In an embodiment, the transistor (indicated with a circle in FIG. 1) of OECT biosensor 30 may be positioned to overlap a given electrode such that a measurement can be performed by OECT biosensor 30 when the liquid volume is moved to a position between said given electrode and transistor.

[0058] The use of a conductive material, which effectively forms an electrode of the second substrate, may enhance movement and manipulation of the liquid volume during operation. Thus, when first portion 21a includes conductive material, it is most beneficial if a substantial portion of electrodes 14 overlaps with first portion 21a and that only a small portion of electrodes 14 is dedicated to a ‘sensing region’ overlapping with second portion 21b.

[0059] Sensing unit 50 may be configured to be coupled to OECT biosensor 30 during operation to initiate a measurement using OECT biosensor 30 and acquire a corresponding measurement result. In particular, sensing unit 50 may apply a gate voltage to the transistor of OECT biosensor 30 and may measure a corresponding (drain-source) current.

[0060] Result deriving unit 60 may be configured to receive the measurement result from sensing unit 50 and may process the measurement result to derive information therefrom and format the information in a manner interpretable by a user. For example, result deriving unit 60 may directly output the measured current for one or multiple gate voltages and / or drain voltages, one or multiple respective liquid volumes, or the like. Alternatively, result deriving unit 60 may derive, based on the measured current, an ion concentration present in the liquid volume.

[0061] It is noted that result deriving unit 60 may be external to DMF system 100. For example, result deriving unit 60 may be implemented as software or hardware in a processor, computer, remote server, or the like, configured to communicate (e.g., by wired or wireless connection) with sensing unit 50 for the purpose of initiating the measurement and receiving the corresponding measurement result. Said processor, computer, remote server, or the like, may additionally oralternatively be configured to communicate (e.g., by wired or wireless connection) with control unit 40 to control DMF system 100 by providing one or more instructions to control unit 40. Although not illustrated in FIG. 1, second substrate 21 may have further sensors arranged (e.g., integrated) thereon in addition to OECT biosensor 30. For example, said further sensor(s) may include one or more electrochemical sensors, such as an electrochemical impedance spectroscopy (EIS) based sensor, a field-effect transistor (FET) based biosensor, or an aptamerbased biosensor potentiostat. Additionally or alternatively, the further sensor(s) may include one or more optical sensors, such as a fiber-tip photonic crystals biosensor, a surface enhanced Raman scattering (SERS) based sensor, or a surface plasmon resonance (SPR) based sensor. Additional sensors and sensor types not mentioned are equally envisaged, as will be appreciated by the skilled person.

[0062] During operation, first substrate unit 10 and second substrate unit 20 are arranged spaced apart and in parallel to each other, as illustrated in a simplified manner in FIG. 1 and described in more detail with reference to FIG. 3.

[0063] FIG. 2 A illustrates a simplified top view of first substrate unit 10 in accordance with an embodiment of the present invention. Since the various elements of first substrate unit 10 have already been described with reference to FIG. 1 , a detailed description thereof is omitted.

[0064] FIG. 2B illustrates a simplified top view of second substrate unit 10 in accordance with an embodiment of the present disclosure. Various elements of second substrate unit 20 have already been described with reference to FIG. 1 , and a detailed description of those elements is therefore omitted.

[0065] In this embodiment, a plurality of OECT biosensors 30-1 through 30-4 are arranged on second substrate 21, in particular on second portion 21b of second substrate 21. Each OECT biosensor 30-1 through 30-4 may include a respective transistor 31-1 through 31-4, terminals of which are coupled (e.g., via wiring arranged on second substrate 21) to respective contacts including a drain contact de, a source contact sc, and a gate contact gc (only indicated for OECT biosensor 30-1).

[0066] The use of multiple OECT biosensors 30-1 through 30-4 enables performing multiple measurements concurrently (e.g., on multiple liquid volumes), or consecutively (e.g., for the same liquid volume) under different circumstances. As described further below with reference to FIG. 5, OECT biosensors 30-1 through 30-4 may have respective sensitivities or ion selectivity, which, through their convenient combined integration into a single DMF system 100, enables a user of DMF system 100 to significantly reduce measurement time and improve overall measurement accuracy.

[0067] FIG. 3 illustrates a simplified cross-sectional view of at least part of DMF system 100, in particular first substrate unit 10 and second substrate unit 20. Various elements of second substrateunit 20 have already been described with reference to FIG. 1 , and a detailed description of those elements is therefore omitted.

[0068] As shown in FIG. 3, electrodes 14 may be arranged facing second substrate unit 20 during operation. Similarly, transistor 31 of OECT biosensor 30 may be arranged facing first substrate unit 10 during operation. Here, transistor 31 includes a source terminal 31s, a drain terminal 31d, and a gate terminal 31g. For example, terminals 31s, 31d, 31g are made of metal, such as gold.

[0069] First substrate unit 10 may further comprise a first hydrophobic layer 13 to enhance mobility of liquid between first substrate unit 10 and second substrate unit 20. Similarly, second substrate unit 20 may further comprise a second hydrophobic layer 23. As shown in FIG. 3, hydrophobic layer 23 may be present in first portion 21a of second substrate 21 and may be at least partially omitted in second portion 21b of second substrate 21, in particular where transistor 31 is arranged.

[0070] First substrate unit 10 may further comprise a dielectric layer 12 covering first substrate 11 and electrodes 14. Dielectric layer 12 may be arranged between first substrate 11 and hydrophobic layer 13 (if present). In an example, dielectric layer 12 may be present to enable manipulation of a liquid volume D (e.g., a (micro)droplet) through electrodes 14 using electrowetting-on-dielectric (EWOD).

[0071] Second substrate unit 20 may further comprise conductive material 22, e.g., a conductive layer. Conductive material 22 may be arranged between second substrate 21 and hydrophobic layer 23 (if present). As shown in FIG. 3, conductive material 22 may be present in first portion 21a of substrate 21 and may be at least partially omitted in second portion 21b of second substrate 21, in particular where transistor 31 is arranged. Preferably, conductive material 22 is electrically grounded during operation. For example, conductive material 22 of second substrate unit 20 and ground electrode 17 of first substrate unit 10 may be electrically biased at the same reference voltage, e.g., ground.

[0072] As shown in FIG. 3, liquid volume D may be present between first substrate unit 10 and second substrate 20 at a first position Pl and may be adjacent to a given electrode 14. To move liquid volume D, an adjacent electrode 14 may be actuated by applying a positive voltage thereto, which attracts liquid volume D to move to said adjacent electrode 14. In this manner, liquid volumes can be manipulated to move to a desired position between first substrate unit 10 and second substrate unit 20.

[0073] The desired position may for example be a second position P2, which is directly adjacent to transistor 31 of OECT biosensor 30. When liquid volume D is moved to second position P2, a measurement may be performed using OECT biosensor 30 by controlling transistor 31 accordingly. In particular, depending on an operating mode of transistor 31 (e.g., a depletion-mode transistor or an accumulation-mode transistor), ions in liquid volume D may be attracted to orrepelled from a channel extending between source terminal 31s and drain terminal 3 Id, thereby changing its conductivity. As a result, after applying a gate voltage, a (drain-source) current can be measured which may be indicative of the contents of liquid volume D.

[0074] At a third position P3, cells in a medium C (e.g., a cell culture) may be provided on second substrate 21. In that case, liquid volume D may first be moved from first position Pl to third position P3, where ion exchange with cell culture C is allowed for a given period of time.

[0075] Afterwards, liquid volume D may be moved from third position P3 to second position P2 for performing the measurement. Accordingly, a measurement can be performed of ion concentrations that are indicative of the contents (e.g., the health) of cell culture C, for example for the purpose of monitoring.

[0076] The following parameters are provided as a suitable example only, and are not intended to limit the present disclosure in any way or form. Various modifications can be made, as will be appreciated by those skilled in the art. A suitable channel size of transistor 31 may be 25 pm x 100 pm, and an area of gate terminal 31g may be 1000 pm2. Electrodes 14 may be patterned in a 2 mm x 2mm configuration. Second substrate unit 20 may be constructed on a 25 mm x 75 mm glass slide and may be coated with ITO only in first portion 21a. A gap between first substrate unit 10 and second substrate unit 20 may be about 120 pm. Depending on the implementation, ion concentrations may be measured using transistor 31 by applying one or more gate voltages ranging from 0 to 600 mV, with a fixed drain voltage, such as 100 mV, and detecting changes in the (drainsource) current. A suitable actuation voltage for electrodes 14 to manipulate and move liquid volume D may be about 120 V. In first portion 21a, an opening of 2 mm x 2 mm in conductive material 22 and hydrophobic layer 23 may be provided to receive a cell culture site, such as cells in medium C of FIG. 3, directly in second substrate 21 (e.g., bare glass).

[0077] Next, a fabrication process of transistor 31 of OECT biosensor 30 will be described step-by-step with reference to FIG. 4A-4L.

[0078] FIG. 4A shows part of second substrate 21, including first portion 21a and second portion 21b. For example, first portion 21a is conductive material 22, such as ITO and second portion 21b is glass. A first mask layer 71 (e.g., a photoresist layer) is applied to second substrate 21, for example by means of a photolithography process. Once applied, first mask layer 71 includes gaps where terminals of transistor 31 are to be formed on second substrate 21, in this case in particular in second portion.

[0079] In FIG. 4B, a metallic layer 72 is deposited, covering first mask layer 71 and a portion of second substrate 21 that is exposed through first mask layer 71. In an example, metallic layer 72 may be made of gold or other suitable metals or metal alloys for terminals or electrodes.In FIG. 4C, first mask layer 71 is lifted off, leaving only portions of metallic layer 72 that was deposited on the exposed portion of second substrate 21, which portions respectively form gate terminal 31g, source terminal 31s, and drain terminal 3 Id of transistor 31.

[0080] In FIG. 4D, a first protective layer 73 is applied, in particular to first portion 21a of second substrate 21. For example, Kapton tape may be used as first protective layer 73, which is intended to cover and protect first portion 21a throughout the fabrication process, since the fabrication process pertains to transistor 31 in second portion 21b rather than first portion 21a of second substrate 21. Throughout the fabrication process, first protective layer 73 may be removed one or multiple times and a new protective layer may be placed in its place to prevent excess buildup of material on said protective layer during the whole fabrication process. However, this removal and reapplication of protective layers is merely an optional step in the fabrication process.

[0081] In FIG. 4E, a first Parylene coating 74 is applied across second substrate 21. The Parylene coating forms a dielectric layer on top of terminals 31g, 31s, and 31d to prevent them from being in direct electrical connection to electrodes 14 during operation (e.g., through the liquid volume). It also acts as a protective layer to shield the underlying metallic contacts from degradation due to exposure to liquid volumes. Alternative dielectric materials may be used instead of first Parylene coating 74, as will be appreciated by the skilled person.

[0082] In FIG. 4F, a sacrificial layer 75 is applied covering first Parylene coating 74, which is a layer that can be easily washed away after performing a particular later fabrication step.

[0083] In FIG. 4G, a second Parylene coating 76 is applied covering sacrificial layer 75.

[0084] In FIG. 4H, a second mask layer 77 (e.g., a photoresist layer) is applied, for example by means of a photolithography process. Once applied, second mask layer 77 includes a first gap corresponding to a position of gate terminal 31g and a second gap corresponding to the channel extending between source terminal 31s and drain terminal 3 Id. Preferably, the second gap is slightly more elongated than a length of the channel, such that it also partially overlaps with source terminal 31s and drain terminal 3 Id.

[0085] Also shown in FIG. 4H is that, as discussed with reference to FIG. 4D, first protective layer 73 has been removed and replaced with a new first protective layer 73’ prior to applying second mask layer 77.

[0086] In FIG. 41, an etching process E is performed to remove material exposed through second mask layer 77. In particular, part of first Parylene coating 74, sacrificial layer 75, and second Parylene coating 76 may be removed by etching process E, thereby exposing gate terminal 31g, part of source terminal 31s and drain terminal 3 Id, and a portion of second substrate 21 overlapping with the channel extending between source terminal 31s and drain terminal 3 Id. Etching process E may for example be a reactive ion etching (RIE) process. In addition, FIG. 41shows that it is possible to remove new first protective layer 73’ from first portion 21a of second substrate 21.

[0087] In FIG. 4J, if protective layer 73 or new first protective layer 73’ have been removed previously, a second protective layer 78 is applied, in particular to first portion 21a of second substrate 21. Second protective layer 78 may be identical or similar to first protective layer 73 or new first protective layer 73’.

[0088] In FIG. 4K, a Poly(3,4-ethylenedioxythiophene):poly(styrenesulfonate) coating 79 (PEDOT:PSS for short) is applied. PEDOT:PSS is a widely used conductive polymer that consists of PEDOT (Poly (3 ,4-ethylenedioxy thiophene)), a conjugated polymer that provides electrical conductivity, and PSS (Poly(styrenesulfonate)), a counterion that makes PEDOT water-dispersible, i.e., processable from aqueous solutions.

[0089] In FIG. 4L, sacrificial layer 75 is lifted off, leaving only parts of first Parylene coating 74, the terminals of transistor 31, PEDOT:PSS coating 79 on second portion 21b of second substrate 21. Additionally, protective layer 78 is removed again, exposing first portion 21a of second substrate 21.

[0090] FIG. 5 illustrates transistor 31 of OECT biosensor 30 according to an embodiment of the present disclosure.

[0091] Further to the process illustrated with reference to FIG. 4A - 4L, transistor 31 may be provided with an ion-selective membrane 32 arranged to partially surround the channel extending between source terminal 31s and drain terminal 3 Id. To that end, OECT biosensor 30 may be considered an ion-sensitive OECT (IS -OECT) biosensor.

[0092] Ion-selective membrane 32 may be substantially impermeable to particular ions while being permeable to one (or more) specific ions. For example, ion-selective membrane 32 may be impermeable By providing an ion-selective membrane, the IS -OECT biosensor may be made more sensitive to certain ions relative to other ions, rendering it possible to measure ion concentrations of said certain ions in the liquid volume with no or only limited influence from other ions.

[0093] In an embodiment, multiple IS-OECT biosensors with different ion-sensitive membranes and thus respectively different ion selectivity may be integrated on second substrate 21. For example, referring to FIG. 2B, OECT biosensor 30-1 may be an IS-OECT biosensor with a first ion-sensitive membrane (e.g., more sensitive to Potassium), and OECT biosensor 30-2 may be an IS-OECT biosensor with a second ion-sensitive membrane (e.g., more sensitive to Sodium). Each of OECT biosensors 30-1 through 30-4 may have different ion-sensitive membranes. Alternatively, at least one of OECT biosensors 30-1 through 30-4 may not have an ion-sensitive membrane.

[0094] In an example implementation of an IS-OECT biosensor, a Poly(sodium 4-styerenesulfonate) (i.e., PSS:Na) layer 33 is provided between PEDOT:PSS coating 79 and ion-selective membrane 32 of transistor 31. This layer is a polyanionic polymer with mobile Na+ions,that may be used in OECTs as an internal ion reservoir to enhance ion transport between ion-selective membrane 32 and the channel of transistor 31 , which for example comprises PEDOT:PSS. Its negatively charged backbone facilitates efficient cation migration under a positive gate bias, improving gating efficiency, response time, and device stability.

[0095] FIG. 6 illustrates a method for operating a DMF system (e.g., DMF system 100 as described above) in accordance with an embodiment of the present disclosure.

[0096] In operation S 1 , the liquid volume is provided between the first and second substrate unit. For example, an amount of liquid is provided on reservoir electrode 15 of first substrate unit 10, the amount of liquid including the liquid volume to be manipulated. Accordingly, the plurality of electrodes 14 may be used (e.g., controlled by control unit 40 of FIG. 1) to acquire (e.g., withdraw) the liquid volume from the amount of liquid on reservoir electrode 15. Electrode 14 directly adjacent to reservoir electrode 15 may be considered to represent a first position of liquid volume.

[0097] In operation S2, the liquid volume is moved, using the plurality of electrodes, from the first position that is away from the OECT biosensor to a second position adjacent to the OECT biosensor in the first direction. That is, the second position may be a position between first substrate unit and second substrate unit that enables the OECT biosensor to perform a measurement on the liquid volume when the liquid volume is present at said second position.

[0098] In an embodiment, the liquid volume may first be moved from the first position to a third position, at which cells are provided in a medium (e.g., a cell culture) between the first and second substrate (e.g., before operation S2, and before, during, or after operation SI). At the third position, the liquid volume may be allowed to exchange ions with the medium including the cells for a given period of time, such that a subsequent measurement using the OECT biosensor is indicative of an ion concentration of the cell culture. After said given period of time, the liquid volume may be moved again using the plurality of electrodes from the third position to the second position adjacent to the OECT biosensor.

[0099] In operation S3, once the liquid volume is at the third position, a measurement is performed using the OECT biosensor. For example, a gate voltage is applied to the OECT biosensor and a corresponding (drain-source) current is measured. This operation may further include processing the current to derive a corresponding ion concentration in the liquid volume, for example the ion concentration of a given ion or an overall ion concentration.

[0100] FIG. 7A is a diagram representing transfer curves of a Na+-selective IS-OECT measured under DMF operation for different concentrations of Sodium Chloride (NaCl) and Potassium Chloride (KC1). The x-axis represents a gate voltage applied to the IS-OECT in Volts, and the y-axis represents a drain current response at given concentrations in Ampere. Solid lines are used to represent transfer curves in response to a NaCl solution, whereas dashed lines are used to represent transfer curves in response to a KC1 solution.For the measurement, electrolyte droplets containing NaCl (primary ion) and KC1 (interfering ion) with concentrations ranging from 105to 101M were sequentially delivered to the OECT gate using electrowetting-based actuation. Measurements were performed at a fixed drain voltage of Vd = -0.1 V while sweeping the gate voltage from 0 to 0.6 V.

[0101] When delivering the KC1 droplets, the OECT channel exhibited only weak and nearly concentration-independent de-doping. Across the entire KC1 concentration range, the modulation of the drain current remained small and rapidly reached a plateau. This indicates minimal interaction between K+-ions and the Na+-selective membrane. Even at the highest KC1 concentration, the induced de-doping remained significantly lower than that obtained with NaCl at lower concentrations.

[0102] In contrast, when NaCl droplets were delivered to the same device, the OECT channel showed a strong and concentration-dependent de-doping response. Increasing the NaCl concentration progressively reduced the drain current, consistent with enhanced Na+-ion transport through the selective membrane and increased ion penetration into the channel. Notably, even the lowest NaCl concentrations produced a larger current modulation than the highest KC1 concentrations, despite the latter being orders of magnitude higher.

[0103] Together, these results demonstrate that the DMF-integrated Na+-selective IS-OECT selectively responds to Nations, while remaining largely insensitive to interfering ions such as K+. The plateaued and limited response to KC1, combined with the response to NaCl, confirms that selective Na+-ion transport dominates the observed channel de-doping, rather than non-specific ionic effects.

[0104] FIG. 7B is a diagram representing drain current (on the y-axis) as a function of electrolyte concentration (on the x-axis) for the Na+-selective IS-OECT, extracted from the transfer curves in FIG. 7A at a fixed gate voltage of vg= 0.6 V and drain voltage of Vd = -0.1 V. The solid line represents a NaCl solution, whereas the dashed line represents a KC1 solution.

[0105] When NaCl droplets were delivered to the Na+-selective IS-OECT, the drain current decreased with increasing concentration, showing a clear, concentration-dependent response and indicating progressively stronger channel de-doping. In contrast, KC1 droplets remained nearly costant and induced only minor, non-systematic variations in the drain current, with no clear dependence on concentration. The markedly different trends observed for NaCl and KC1 further confirm the selectivity towards Nations over K+-ions.

[0106] FIG. 8 A shows transfer curves for a K+-selective IS-OECT measured under DMF operation for different concentrations (in M) of NaCl and KC1. The x-axis represents a gate voltage applied to the IS-OECT in Volts, and the y-axis represents a drain current response at given concentrations in Ampere. Solid lines are used to represent transfer curves in response to a NaCl solution, whereas dashed lines are used to represent transfer curves in response to a KC1 solution.For this measurement, electrolyte droplets containing NaCl (primary ion) and KC1 (interfering ion) with concentrations ranging from 104to 101M were sequentially delivered to the OECT gate using electrowetting-based actuation. Measurements were performed at a fixed drain voltage of Vd = -0.1 V while sweeping the gate voltage from 0 to 0.6 V.

[0107] When delivering the NaCl droplets, the OECT channel showed only a limited de-doping response across the investigated concentration range. In contrast, when delivering KC1 droplets, the channel exhibited a concentration-dependent de-doping, with increasing KC1 concentration leading to a progressively larger decrease in the drain current. At identical concentrations, KC1 consistently induced a larger modulation of the drain current than NaCl, indicating preferential K+transport through the selective membrane.

[0108] These results demonstrate that the DMF-integrated OECT selectively responds to K+ions, while remaining largely insensitive to Na+, and confirm that the observed channel modulation originates from selective ion transport rather than non-specific electrolyte effects.

[0109] FIG. 8B is a diagram representing drain current (on the y-axis) as a function of electrolyte concentration (on the x-axis) for the K+-selective IS -OECT, extracted from the transfer curves in FIG. 8 A at a fixed gate voltage of vg= 0.6 V and drain voltage of Vd = -0.1 V. The solid line represents a NaCl solution, whereas the dashed line represents a KC1 solution.

[0110] When NaCl droplets were delivered to the device, the drain current showed only small variations with concentration, indicating a weak and non-systematic de-doping response. In contrast, when KC1 droplets were delivered, the drain current changed strongly with increasing concentration, which reflects enhanced channel de-doping driven by selective K+transport.

[0111] The significantly larger current modulation observed for KC1 compared to NaCl at the same concentrations confirms that the device responds preferentially to K+-ions, while remaining largely insensitive to Na+-ions. These results further validate the high ion selectivity of the K+-selective IS-OECT under DMF operation.

[0112] With reference to the measurement results shown in FIG. 7A, 7B, 8A, and 8B, the ion-selective membrane (ISM) used in the tested device comprises a dense, plasticized poly( vinyl chloride) (PVC) matrix incorporating a selective ionophore and, where applicable, a lipophilic ionic additive. Ion selectivity using the ISM is achieved through ionophore-mediated complexation and partitioning of target ions within the hydrophobic polymer matrix.

[0113] Referring to FIG. 7A and 7B, for the Na+-selective membrane, the composition includes 26.8 wt% high molecular weight PVC, 6.5 wt% sodium Ionophore X, and 66.7 wt% 2-nitrophenyl octyl ether as plasticizer, dissolved in THF (770 mg in 11.7 mL). This composition is, however, merely exemplary and other ratios and / or compositions to achieve Na+-selectivity may be equally envisaged.Referring to FIG. 8 A and 8B, for the K+-selective membrane, the composition consists of 36.5 wt% high molecular weight PVC, 2.5 wt% potassium Ionophore III, 0.5 wt% potassium tetrakis(4-chlorophenyl)borate as lipophilic additive, and 60.5 wt% diisodecyl adipate as plasticizer, dissolved in THF (500 mg in 5 mL). This composition is, however, merely exemplary and other ratios and / or compositions to achieve K+-sensitivity may be equally envisaged.

[0114] The ISM solutions were drop-cast onto the device and dried overnight at room temperature, forming dense, non-porous membranes with an estimated thickness in the range of approximately 10-50 pm.

[0115] The (IS-)OECT may comprise a PEDOT:PSS channel (Clevios PH1000) formulated with ethylene glycol (conductivity enhancer), DBSA (film-forming additive), and GOPS (crosslinking and adhesion promoter). The channel may be patterned between gold source and drain electrodes (e.g., 100 nm Au with 5 nm Cr adhesion layer). The planar gold gate electrode may be fabricated on the same substrate.

[0116] A crosslinked PSSNa layer (e.g., 1.2% w / v aqueous solution containing DBSA, GOPS, and HC1) may be deposited over the channel region and thermally treated to serve as a solid inner electrolyte reservoir. The ISM may be subsequently deposited on top of this PSSNa layer.

[0117] The device can operate in a planar sandwich configuration comprising: PEDOT:PSS / PSSNa / ISM / liquid electrolyte / Au gate. This architecture enables selective ion-to-electronic transduction through modulation of the PEDOT:PSS channel conductivity in response to ion activity in the external electrolyte.

[0118] In the above description, the present disclosure has been explained using detailed embodiments thereof. However, the present disclosure is not limited to any of these embodiments in particular, and various modifications can be implemented without deviating from the scope of the present disclosure as defined by the appended claims and, in some jurisdictions, their equivalents.

Claims

CLAIMS1. A digital microfluidics, ‘DMF’, system (100), comprising:a first substrate unit (10) comprising a first substrate (11) having arranged thereon a plurality of electrodes (14) for manipulation of a liquid volume (D); anda second substrate unit (20) spaced apart from the first substrate unit (10) in a first direction, wherein the second substrate unit (20) comprises a second substrate (21) having arranged thereon an organic electrochemical transistor, ‘OECT’, biosensor (30).

2. The DMF system (100) according to claim 1, wherein, viewed in the first direction, a position of a transistor (31) of the OECT biosensor (30) overlaps with at least one of the plurality of electrodes (14).

3. The DMF system (100) according to claim 1 or 2, wherein the second substrate (21) comprises a first portion (21a) and a second portion (21b), wherein the first portion (21a) and the second portion (21b) each overlap with at least some of the plurality of electrodes, wherein the first portion (21a) includes conductive material (22).

4. The DMF system (100) according to claim 3, wherein the OECT biosensor (30) is arranged on the second portion (21b).

5. The DMF system (100) according to claim 3 or 4, wherein the first portion (21a) is coated with the conductive material (22).

6. The DMF system (100) according to any of the claims 3-5, wherein the conductive material (22) comprises indium tin oxide, ‘ITO’.

7. The DMF system (100) according to any of the claims 3-6, wherein the second substrate (21) is made of glass.

8. The DMF system (100) according to any of the claims 3-7, wherein the conductive material (22) is configured to be electrically grounded during operation.

9. The DMF system (100) according to any of the previous claims, wherein the first substrate (11) is coated with a first hydrophobic layer (13) facing the second substrate (21), and / orwherein the second substrate (21) is coated with a second hydrophobic layer (23) facing the first substrate (11).

10. The DMF system (100) according to claim 9, wherein the first substrate unit (10) further comprises a dielectric layer (12) between the first hydrophobic layer (13) and the first substrate (11).

11. The DMF system (100) according to claim 10, wherein the DMF system (100) is configured to manipulate the liquid volume (D) using electro wetting-on-dielec trie, ‘EWOD’.

12. The DMF system (100) according to any of the previous claims, wherein the plurality of electrodes (14) are integrated on the first substrate (11), and / or wherein the transistor (31) of the OECT biosensor (30) is integrated on the second substrate (21).

13. The DMF system (100) according to claim 12, wherein the transistor (31) of the OECT biosensor (30) comprises a source terminal (31s), a drain terminal (31d), and a gate terminal (31g) that are arranged in-plane.

14. The DMF system (100) according to any of the previous claims, wherein the OECT biosensor (30) is an ion-selective OECT, ‘IS-OECT’, biosensor including an ion-selective membrane (32).

15. The DMF system (100) according to claims 13 and 14, wherein the ion-selective membrane is disposed over a channel of the transistor (31) that extends between the source terminal (31s) and the drain terminal (3 Id).

16. The DMF system (100) according to any of the previous claims, comprising a plurality of said OECT biosensors (30-1, 30-2, 30-3, 30-4).

17. The DMF system (100) according to claims 14 and 16, wherein the plurality of IS-OECT biosensors (30-1, 30-2, 30-3, 30-4) comprises a first IS-OECT biosensor with a first ion-selective membrane, and a second IS-OECT biosensor with a second ion-selective membrane, wherein the first ion-selective membrane and the second ion-selective membrane have different ion selectivity.

18. The DMF system (100) according to any of the previous claims, wherein the plurality of electrodes (14) is arranged in a matrix of rows and columns.

19. The DMF system (100) according to any of the previous claims, wherein the first substrate unit (10) further comprises at least one reservoir electrode (15) configured to hold an amount of liquid from which the liquid volume (D) to be manipulated can be drawn by the plurality of electrodes (14),wherein, preferably, a size of the at least one reservoir electrode (15) is greater than a size of each of the plurality of electrodes (14).

20. The DMF system (100) according to any of the previous claims, further comprising a control unit (40) configured to actuate the plurality of electrodes (14) in a given manner to move the liquid volume (D) to a desired location between the first and second substrate unit (10, 20).

21. The DMF system (100) according to any of the previous claims, further comprising a sensing unit (50) configured to apply a gate voltage to the OECT biosensor (30) and sense a corresponding current through the OECT biosensor (30).

22. The DMF system (100) according to claim 21, further comprising a result deriving unit (60) configured to determine a characteristic of the liquid volume (D) based on the current through the OECT biosensor (30) received from the sensing unit (50),wherein the characteristic preferably includes an ion concentration present in the liquid volume (D), more preferably of a given ion, such as Sodium, Potassium, and / or a pH level.

23. The DMF system (100) according to any of the previous claims, wherein the first substrate unit (10) and the second substrate unit (20) are formed as plates that extend in parallel to each other.

24. The DMF system (100) according to any of the previous claims, further comprising at least one further sensor at least partially integrated on the second substrate (21), wherein the at least one further sensor preferably includes:one or more electrochemical sensors, such as an electrochemical impedance spectroscopy, ‘EIS’, based sensor, a field-effect transistor, ‘FET’, -biosensor, or an aptamer-based biosensor potentiostat; and / orone or more optical sensors, such as a fiber-tip photonic crystals biosensor, a surface enhanced Raman scattering, ‘SERS’, sensor, or a surface plasmon resonance, ‘SPR’, sensor.

25. A wearable electronic device comprising the DMF system (100) as defined in any of the previous claims.

26. An assembly, comprising:a first substrate unit (10) comprising a first substrate (11) having arranged thereon a plurality of electrodes (12) for digital microfluidics, ‘DMF’, based manipulation of a liquid volume (D); anda second substrate unit (20) comprising a second substrate (21) having arranged thereon an organic electrochemical transistor, ‘OECT’, biosensor (30).

27. A method for operating the DMF system as defined in any of the claims 1-24, comprising:a) providing (SI) the liquid volume between the first and second substrate unit;b) moving (S2), using the plurality of electrodes, the liquid volume from a first position that is away from the OECT biosensor to a second position adjacent to the OECT biosensor in the first direction; andc) performing (S3) a measurement using the OECT biosensor.

28. The method according to claim 27, wherein step a) further comprises providing cells in a medium between the first and second substrate at a third position different from the first and second position,wherein, step b) comprises:moving, using the plurality of electrodes, the liquid volume from the first position to the third position;allowing the cells in the medium to exchange ions with the liquid volume; and moving, using the plurality of electrodes, the liquid volume from the third position to the second position.

29. The method according to claim 27 or 28, wherein step a) comprises: providing an amount of liquid on a reservoir electrode of the first substrate unit, the amount of liquid including the liquid volume; andacquiring, using the plurality of electrodes, the liquid volume from the amount of liquid to provide the liquid volume at the first position.

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