Digital microfluidic system and method of dispensing a droplet on the same

Abstract

This invention discloses a digital microfluidic system comprising: (a) a base plate including an electrode array comprising a plurality of digital microfluidic propulsion electrodes; (b) a top plate including a common top electrode; (c) a controller coupled to the processing unit, the common top electrode, and the bottom electrode array; and (d) a processing unit operably programmable for: receiving input commands relating to droplet diameter and aspect ratio; calculating actuation parameters including the length of an actuator frame, the length of an actuator neck, and the height of an actuator head for dispensing a droplet having the diameter and aspect ratio of the input commands; and outputting electrode actuation to the controller, the electrode actuation commands relating to implementing a dispensing drive sequence for calculating the actuation parameters for dispensing a droplet having the input diameter and aspect ratio; wherein the electrodes have a dimension smaller than the droplet diameter.

Description

Technical Field

[0001] This invention relates to a method for applying droplets to a digital microfluidic system and a digital microfluidic system. Prior Technology

[0002] Digital microfluidic devices utilize individual electrodes to propel, split, and join droplets in confined environments, thereby providing a "lab-on-a-chip" experience. Digital microfluidic devices are alternatively referred to as electron wetting over dielectrics, or "EWoD," to compete with microfluidic systems that rely on electrophoretic flow rates and / or micropumps. A 2012 review of electron wetting technology by Wheeler in "Digital Microfluidics," Annu. Rev. Anal. Chem. 2012, 5:413-40, is incorporated herein by reference in its entirety. This technology allows for sample preparation, analysis, and synthetic chemistry using minute quantities of samples and reagents. In recent years, controlled droplet manipulation within microfluidic cells using electron wetting has become commercially viable, and products are now available from larger life science companies such as Oxford Nanopore.

[0003] Most literature on EWoDs involves so-called "passive matrix" devices (also known as "segmented" devices), which directly drive ten to twenty electrodes with a controller. While segmented devices are easy to manufacture, the number of electrodes is limited by space and driving constraints. Therefore, large-scale parallel analysis and reactions are not possible in passive matrix devices. In contrast, "active matrix" devices (also known as active matrix EWoDs or AM-EWoDs) can have thousands, hundreds of thousands, or even millions of addressable electrodes. Electrodes are typically switched by thin-film transistors (TFTs), and droplet movement is programmable, allowing AM-EWoD arrays to be used as general-purpose devices, enabling a high degree of freedom in controlling multiple droplets and simultaneously performing analytical methods.

[0004] Digital microfluidic systems are designed for biological or chemical applications. These often require large volumes of liquid to be introduced into the device via reservoirs, followed by dispensing in smaller quantities for reactions or other functions. Traditionally, dispensing is achieved using reservoirs with large segments, followed by a series of steps to dispense droplets onto a single-size track. The basic procedure for dispensing typically begins by extending a line of liquid from the reservoir. A thin neck is then formed between the reservoir and the initial droplet, and the reservoir and droplet move in opposite directions. This approach is applicable but often suffers from reproducibility problems due to large variations in reservoir volume and is limited to dispensing only single-size droplets due to the architecture of the rest of the array. For example, International Publication WO 2008 / 124846 describes a general method for stretching a drop of fluid to a necked portion and subsequently breaking up daughter droplets. Its system relies on a segmented array where there is no choice in the size of the resulting droplets. Multi-segmented structures are used for the reservoir region, but only one wide channel is used for droplet dispensing. Nikapitiya et al. (Micro and Nano Syst Lett (2017) 5:24) developed a method using a specific structure to achieve a coefficient of variation (CV) of less than 1%. The novelty lies in how the neck-shaped portion is formed and how the droplet is disintegrated (along the diagonal), thereby producing cleaner and reproducible symmetry for disintegration. However, this design is segmented and limited to a fixed droplet size.

[0005] Cho et al. (Journal of Microelectromechanical Systems, Vol. 12, No. 1, February 2003) provide a physical analysis of how basic droplet operations are performed on an electronic wetting device and use physical parameters of the electronic wetting system, such as dielectric constant, voltage, and thickness, to define the parameters that need to be adjusted to maximize the efficiency of each operation. Specifically, the references describe the requirements for the formation of neck-like portions relative to splitting electrodes and multiple parameters. U.S. Patent No. 8,936,708 describes a method for smaller droplets to split from larger droplets. The references primarily deal with defining prototypes of pixels with different geometries (e.g., hexagons) and how droplets are split throughout such pixels. However, no precise method is provided for systematically dispensing droplets of different sizes. U.S. Patent No. 8,834,695 discusses the possibility of using small electrodes to dispense larger models that can be used as dispensing reservoirs. Methods for size control utilize the aggregation of small droplets into larger droplets, but do not provide systems with droplets of variable size or effective application, nor do they focus on methods for improving CV. Summary of the Invention

[0006] In the first embodiment, this application addresses the shortcomings of prior art by providing an interleaved method for dispensing droplets on a digital microfluidic system, the system comprising: (a) a substrate including: a bottom electrode array including a plurality of digital microfluidic propulsion electrodes; and a first dielectric layer covering the bottom electrode array; (b) a top plate including: a common top electrode; and a second dielectric layer covering the common top electrode; (c) a processing unit operably programmable for performing a microfluidic actuation method; and; and (d) a controller operably coupled to the processing unit, the common top electrode, and the bottom electrode array, wherein the controller is configured to provide a propulsion voltage between the common top electrode and the substrate propulsion electrode. The method includes: receiving input instructions in a processing unit, the input instructions relating to droplet diameter and aspect ratio; calculating in the processing unit actuation parameters including the length of an actuator frame, the length of an actuator neck, and the height of an actuator head, for dispensing a droplet having the diameter and aspect ratio of the input instructions; outputting electrode actuation instructions from the processing unit to a controller, the electrode actuation instructions relating to a dispensing drive sequence for implementing the calculated actuation parameters; executing the dispensing drive sequence on a propulsion electrode to: shape fluid in a reservoir to form an actuator frame and an actuator neck; disintegrate the droplet from the head of the neck; and return the neck fluid to the reservoir, wherein the electrode has a dimension smaller than the droplet diameter.

[0007] In a second embodiment, this application provides a novel digital microfluidic system comprising: (a) a substrate including: a bottom electrode array including a plurality of digital microfluidic propulsion electrodes; and a first dielectric layer covering the bottom electrode array; (b) a top plate including: a common top electrode; and a second dielectric layer covering the common top electrode; (c) a processing unit; and (d) a controller operatively coupled to the processing unit, the common top electrode, and the bottom electrode array, wherein the controller is configured to provide a propulsion voltage between the common top electrode and the substrate propulsion electrodes. The processing unit is operably programmed to: receive input instructions relating to droplet diameter and aspect ratio; calculate actuation parameters including the length of the actuator frame, the length of the actuator neck, and the height of the actuator head, in order to apply a droplet having the diameter and aspect ratio of the input instructions; and output electrode actuation to the controller, the electrode actuation instructions relating to an application drive sequence for implementing the calculated actuation parameters, in order to apply a droplet having the input diameter and aspect ratio; wherein the electrodes have a dimension smaller than the droplet diameter.

[0008] In the third state, this paper provides an improved method for dispensing droplets on a digital microfluidic system. The method includes extending a liquid line from a reservoir, forming an actuation neck between the reservoir and an initial droplet, and dissolving the droplet from an actuation head at the neck. The improvement includes raising the height of the actuation head to a forward dissolution height before the droplet dissolves from the head. Simple Explanation of the Diagram

[0009] Figure 1 shows a conventional microfluidic device including a common top electrode.

[0010] Figure 2 is a schematic diagram of the TFT architecture of multiple drive electrodes used in an EWoD device.

[0011] Figure 3 is a schematic diagram of a portion of the base TFT array, including the push electrode, thin film transistor, storage capacitor, dielectric layer and hydrophobic layer.

[0012] Figure 4 is a schematic top view of a reservoir defined by a high-density electrode grid.

[0013] Figure 5 is a top view of the reservoir in Figure 4, where the electrode grid is not shown for clarity. Figures 5A and 5B illustrate the actuator neck at different heights.

[0014] Figure 6 is a top view of the reservoir in Figure 4, in which the actuation parameters used to implement the application drive sequence are identified.

[0015] Figure 7 is a flowchart illustrating an exemplary droplet application method according to this application.

[0016] Figure 8 is a schematic diagram of the droplet application model.

[0017] Figure 9 schematically illustrates the operation of fluid in the collection reservoir.

[0018] Figure 10 illustrates the formation of the frame and neck.

[0019] Figure 11 illustrates the disintegration of the droplet from the neck.

[0020] Figure 12A illustrates the changes in droplet decomposition, including the formation of an elongated "timing neck". Figure 12B shows the effect of the timing neck on the negative radius of curvature of the pinch point.

[0021] Figure 13A shows the variation of droplet decomposition, where the head height increases to a larger, later head height. Figure 13B illustrates the effect of the forward head height on the radius of curvature of the pinch point.

[0022] Figure 14 illustrates the mechanism of self-actuated neck cutting of droplets.

[0023] Figure 15 illustrates the voltage models on the active pixel electrode (Figure 15A) and the non-active pixel electrode (Figure 15B). Implementation

[0024] [definition] Unless otherwise specified, the following terms have their designated meanings.

[0025] The term "actuation" in relation to one or more electrodes means achieving a change in the electrical state of one or more electrodes, which causes the droplet to operate in the presence of the droplet.

[0026] "Droplet" refers to a liquid of a certain volume that is electron-wetting hydrophobic surface and is at least partially defined by a carrier fluid. For example, a droplet may be completely surrounded by a carrier fluid or may be defined by a carrier fluid and one or more surfaces of an EWoD device. Droplets can take on a wide variety of shapes; non-limiting examples include generally disk-shaped, block-shaped, truncated spheres, ellipsoids, spheres, partially compressed spheres, hemispheres, ovoids, cylinders, and various shapes formed during droplet operation, such as merging or splitting, or due to contact between such shapes and one or more working surfaces of the EWoD device. Droplets may include typically polar fluids, such as water, as well as aqueous or anhydrous compositions, or may be mixtures or emulsions comprising aqueous and anhydrous components. Specific compositions of droplets are not particularly relevant, limited only by their electron-wetting hydrophobic working surfaces. In various embodiments, the droplets may include biological samples such as whole blood, lymph, serum, plasma, sweat, tears, saliva, sputum, cerebrospinal fluid, amniotic fluid, semen, vaginal secretions, serous fluid, synovial fluid, pericardial fluid, peritoneal fluid, pleural fluid, exudate, secretions, cystic fluid, bile, urine, gastric juice, intestinal juice, fecal samples, liquids containing single or multiple cells, liquids containing organelles, fluidized tissues, fluidized organisms, liquids containing multicellular organisms, biological swabs, and biological wash solutions. Furthermore, the droplets may include one or more reagents, such as water, deionized water, physiological saline solutions, acidic solutions, alkaline solutions, cleaning solutions, and / or buffer solutions. Other examples of droplet content include reagents such as those used in biochemical protocols, nucleic acid amplification protocols, affinity-based analytical protocols, enzyme analytical protocols, gene sequencing protocols, protein sequencing protocols, and / or protocols for biofluidic analysis. Other examples of reagents include those used in biochemical synthesis methods, such as reagents for the discovery of oligonucleotides in molecular biology and medicine and / or in more than one nucleic acid molecule. Oligonucleotides may contain natural or chemically modified bases and are most commonly used as antisense oligonucleotides, small interfering therapeutic RNA (siRNA) and their bioactive conjugates, primers for DNA sequencing and amplification, probes for detecting complementary DNA or RNA via molecular hybridization, for targeted introduction of mutations and restriction sites (in the case of gene editing techniques, such as CRISPR-Cas9), and tools for synthesizing artificial genes by "synthesizing and stitching together" DNA fragments.

[0027] "Droplet manipulation" means any operation of one or more droplets on a microfluidic device. Droplet manipulation may include, for example: loading droplets into a microfluidic device; dispensing one or more droplets from a source; splitting, separating, or dividing a droplet into two or more droplets; transporting a droplet from one location to another in any direction; merging or combining two or more droplets into a single droplet; diluting droplets; mixing droplets; agitating droplets; deforming droplets; holding droplets in situ; cultivating droplets; heating droplets; vaporizing droplets; cooling droplets; placing droplets; outputting droplets from a microfluidic device; other droplet manipulations described herein; and / or any combination thereof. The terms "merge," "merging," "combine," "combining," and the like are used to describe the formation of a single droplet from two or more droplets. It should be understood that when such terms are used to refer to two or more droplets, any combination of droplet operations sufficient to result in the combination of two or more droplets into one droplet can be used. For example, "merging droplet A with droplet B" can be achieved by bringing droplet A into contact with stationary droplet B, bringing droplet B into contact with stationary droplet A, or bringing droplets A and B into contact with each other. The terms "split," "separate," and "divide" are not intended to imply any particular result regarding the volume of the resulting droplets (i.e., the resulting droplet volumes may be the same or different) or the number of resulting droplets (the number of resulting droplets may be 2, 3, 4, 5, or more). The term "mixing" refers to a droplet operation that results in a more uniform distribution of one or more components within a droplet. Examples of "loading" droplet operations include microdialysis loading, pressure-assisted loading, automated loading, passive loading, and pipette loading. Droplet operations can be electrode-mediated. In some cases, droplet manipulation is further facilitated by the use of hydrophilic and / or hydrophobic regions on the surface and / or by physical barriers.

[0028] When using a reference droplet, the "diameter" is intended to identify the longest straight line segment between two points on the droplet's surface.

[0029] A "gate driver" is a power amplifier that receives a low-power input from a controller (such as a microcontroller integrated circuit (IC)) and generates a large current drive input for the gate of a high-power transistor (such as a TFT). A "source driver" is a power amplifier that generates a large current drive input for the source of a high-power transistor. A "top electrode driver" is a power amplifier that generates a drive input for the top planar electrode of an EWoD device.

[0030] "Nucleic acid molecules" is a general term for single-stranded or double-stranded, sense or antisense DNA or RNA. These molecules are composed of nucleotides, which are monomers consisting of three parts: a 5-carbon sugar, a phosphate group, and a nitrogenous base. If the sugar is ribose, the polymer is RNA (ribonucleic acid); if the sugar is derived from deoxyribose, the polymer is DNA (deoxyribonucleic acid). Nucleic acid molecules vary in length, ranging from oligonucleotides of about 10 to 25 nucleotides commonly used for gene testing, research, and identification to relatively long or very long prokaryotic and eukaryotic genes with sequences of about 1,000, 10,000, or more nucleotides. Their nucleotide residues can be entirely naturally occurring or at least partially chemically modified to, for example, slow down degradation in vivo. Modification of the molecular backbone can be achieved, for example, by introducing nucleoside organothiophosphate (PS) nucleotide residues. Another modification of nucleic acid molecules suitable for medical applications is 2' sugar modification. Modifying the 2'-position sugar enhances the effectiveness of therapeutic oligonucleotides, particularly in antisense oligonucleotide therapy, by increasing their target binding capacity. The two most commonly used modifications are 2'-O-methyl and 2'-fluorine.

[0031] When a liquid in any form (e.g., droplets or a continuous body, whether moving or stationary) is described as being "above," "at," or "on" an electrode, array, matrix, or surface, such liquid may be in direct contact with the electrode / array / matrix / surface or may be in contact with one or more layers or films interposed between the liquid and the electrode / array / matrix / surface.

[0032] When a droplet is described as being "on" or "mounted on" a microfluidic device, it should be understood that the droplet is configured on the device in a manner that facilitates the use of the device to perform one or more droplet operations on the droplet, the droplet is configured on the device in a manner that facilitates the sensing of droplet characteristics or signals, and / or the droplet has been operated on a droplet actuator.

[0033] When referring to multiple items, "each" is intended to identify an individual item in the set, but does not necessarily refer to every item in the set. Exceptions may exist if the explicit content or context clearly specifies otherwise.

[0034] Throughout this specification, references to "one embodiment," "some embodiments," "one or more embodiments," or "embodiment," regardless of whether the term "illustrative" or "non-exclusive" precedes the term "embodiment," mean that a particular feature, structure, material, step, or characteristic described in connection with an embodiment is included in at least one embodiment of the invention. Therefore, the appearance of phrases throughout different places in this specification, such as "in one or more embodiments," "in some embodiments," "in one embodiment," or "in an embodiment," does not necessarily refer to the same embodiment of the invention. Furthermore, in one or more embodiments, a particular feature, structure, material, step, or characteristic may be incorporated in any suitable manner.

[0035] In the context of a microfluidic device, the use of "top" and "bottom" is merely a rule that the two plates can be interchanged, and the device can be oriented in various ways. For example, the top and bottom plates can be generally parallel, while the overall device can be oriented such that the plates are perpendicular to the worktable surface (as opposed to being parallel to the worktable surface shown in the figure). The top or bottom plate may include additional functionality, such as heating via commercially available microheaters and thermocouples integrated with the microfluidic platform and / or temperature sensing.

[0036] This will greatly benefit the precise control of fluid volume for efficient and varied droplet distribution. This capability will also enable complex droplet operations involving multiple droplet-containing reactants, often combined in processes with parallel reactions. Furthermore, importantly, reproducibility is high and size variations remain minimal across all droplet sizes. The liquid can also exhibit varying viscosities and variable surface tension, which can greatly benefit from a highly tunable distribution model. This invention provides a method for distributing droplets of different sizes with high accuracy and reproducibility using a high-density electrode system, such as a thin electrode transistor (TFT) array. Importantly, this robust distribution strategy is applicable to reservoirs that can cover a range of droplet volumes, particularly down to extremely small droplets.

[0037] In some cases, the basic procedure for dispensing remains similar to that reported in the literature, as discussed in the following background: First, the liquid line extends from the reservoir. Then, a thin neck is formed between the reservoir and the initial droplet, and the reservoir and droplet move in opposite directions. Conventional approaches are largely based on segmented arrays with limited control over the dispensing volume and volumetric velocity (CV). This achieves limited control over the reservoir fluid due to the low-density reservoir electrodes. It is also limited in the ability to control the necking characteristics in more than one dimension, as the electrode size is approximately the droplet diameter. Therefore, the ability to dispense fluids of different droplet sizes and viscosities is limited.

[0038] In contrast, this application defines a reservoir and dispensing model that relies on multiple actuation parameters, which can be dynamically adjusted based on variables such as droplet size, viscosity, and surface tension. The model relies on a high-density electrode array, thus eliminating problems typically associated with fixed segmented structures and ensuring uniformity of dispensing across multiple droplet sizes, while allowing for dynamic consideration of residual liquid in the reservoir. This allows for reservoir and neck shaping to define the desired droplet size and achieves clean dispensing with higher accuracy and reproducibility. After neck formation, several strategies can be used for dissolution, depending on droplet characteristics.

[0039] The dispensing method described in this application reduces the failure rate of multi-step droplet operations, such as in complex analyses, thereby improving the reliability of the EWoD microfluidic sleeve. The range of reagents usable on digital microfluidic devices is also expanded, thus improving the scope of feasible applications. It also ensures higher reproducibility of parallel analyses performed at various volume ratios, improving the parallelization capability of the device, especially at lower liquid volumes.

[0040] In a representative embodiment, the substrate of the microfluidic device comprises an active matrix of electron-wetting dielectric (AM-EWoD) arrays having a plurality of elements, each array element including a drive electrode, but also encompassing other configurations for driving the substrate electrodes. The AM-EWoD substrate may be in the form of a transistor active matrix backplane, such as a thin-film transistor (TFT) backplane, wherein each drive electrode is operatively attached to a transistor and capacitor that actively maintain the electrode state, while electrodes of other array elements are addressed. The top electrode circuitry can independently drive the top electrode.

[0041] The propulsion voltage can be defined by the voltage difference between the array electrodes and the top electrode throughout the microfluidic region. By adjusting the frequency and amplitude of the signals driving the array electrodes and the top electrode, the propulsion voltage of each pixel in the array can be controlled to operate the AM-EWoD device in different operating modes according to the different droplet manipulation operations to be performed. In one embodiment, the TFT array can be implemented using amorphous silicon (a-Si), thereby reducing production costs to the point where a single-use device is possible.

[0042] The basic operation of a typical EWoD device is illustrated in the cross-sectional view of Figure 1. EWoD

[0100] Including fluids filled with fillers

[0102] and at least one aqueous droplet

[0104] The microfluidic region. Typically, nonpolar filler fluids are used for manipulating aqueous droplets. Nonpolar fluids can be hydrocarbons, such as dodecane, polysiloxane, or other nonpolar long-chain organic fluids. The gap in the microfluidic region depends on the size of the droplet being processed and is typically in the range of 50 to 200 µm, but can be larger. In the basic configuration of Figure 1, multiple propulsion electrodes...

[0105] Mounted on a single substrate and sharing a common top electrode

[0106] It is placed on the opposing surface. The device further includes a hydrophobic coating on the surface in contact with the oil layer.

[0107] and propulsion electrodes

[0105] with hydrophobic coating

[0107] Dielectric layers between

[0108] (The upper substrate may also include a dielectric layer, but it is not shown in Figure 1). The hydrophobic layer prevents droplets from wetting the surface. When no voltage difference is applied between adjacent electrodes, the droplets will maintain a spherical shape to minimize contact with the hydrophobic surfaces (oil layer and hydrophobic layer). Because the droplets do not wet the surface, they are unlikely to contaminate the surface or interact with other droplets, except when such behavior is required.

[0043] While monolayers with dielectric and hydrophobic properties are possible, such layers typically require a thick inorganic layer with a resulting low dielectric constant (to prevent pinholes), where droplet movement requires over 100 V. For low-voltage propulsion, a thin, pinhole-free inorganic layer with high capacitance is often preferred, topped with a thin organic hydrophobic layer. With this combination, electronic wetting operation with voltages in the range of + / -10 to + / -50 V is possible, within the range available for conventional TFT arrays.

[0044] A hydrophobic layer can be made by depositing a hydrophobic material onto a surface (via a suitable technique) to form a coating. Depending on the hydrophobic material to be applied, exemplary deposition techniques include spin coating, molecular vapor deposition, and chemical vapor deposition. The hydrophobic layer may be more or less wettable, as typically defined by its individual contact angles. Unless otherwise stated, angles are measured herein in degrees (°) or radians (rad), depending on the context. For the purpose of measuring surface hydrophobicity, the term "contact angle" should be understood to refer to the contact angle of the surface relative to deionized (DI) water. A surface is classified as hydrophilic if water has a contact angle between 0° < θ < 90°, and a surface producing a contact angle between 90° < θ < 180° is considered hydrophobic. Typically, a medium contact angle is considered to fall within the range of about 90° to about 120°, while a high contact angle is typically considered to fall within the range of about 120° to about 150°. When the contact angle is 150° < θ, the surface is typically known to be superhydrophobic or ultrahydrophobic. Surface wettability can be measured using analytical methods well-known in this art, such as by applying droplets to the surface and measuring the contact angle using a contact angle goniometer. Anisotropic hydrophobicity can be tested by tilting a substrate with gradient surface wettability along the transverse axis of the model and examining the minimum tilt angle of the movable droplet.

[0045] Hydrophobic layers with medium contact angles typically include one or a blend of fluoropolymers such as PTFE (polytetrafluoroethylene), FEP (fluorinated ethylene propylene), PVF (polyvinylidene fluoride), PVDF (polyvinylidene fluoride), PCTFE (polychlorotrifluoroethylene), PFA (perfluoroalkoxy polymer), FEP (fluorinated ethylene propylene), ETFE (polyethylene tetrafluoroethylene), and ECTFE (polyvinyl chloride trifluoroethylene). Commercially available fluoropolymers include Cytop® (AGC Chemicals, Exton, PA), Teflon® AF (Chemours, Wilmington, DE), and Cytonix's FluoroPel™ coating (Beltsville, MD). The advantage of fluoropolymer films is their high inertness and ability to maintain hydrophobicity, even after exposure to oxidative treatments such as corona treatment and plasma oxidation.

[0046] When a voltage difference is applied between adjacent electrodes, the voltage on one electrode attracts the dielectric to the opposite charge in the droplet at the droplet interface, and the droplet moves toward this electrode, as illustrated in Figure 1. The acceptable voltage required for droplet propulsion depends on the characteristics of the dielectric and hydrophobic layers. AC drive is used to reduce the degradation of the droplet, dielectric, and electrodes through different electrochemical properties. The operating frequency of EWoD is in the range of 100 Hz to 1 MHz, but lower frequencies of 1 kHz or lower are preferred for use with TFTs with limited operating speeds.

[0047] Returning to Figure 1, top electrode

[0106] A single conductive layer, typically set to zero volts or a common voltage value (VCOM), is considered for advancing the electrode.

[0105] The offset voltage is attributed to the capacitor backlash of the TFT used for switching the voltage on the electrode (see Figure 3). The top electrode may also have a square wave applied to increase the voltage throughout the liquid. This configuration allows for a lower push voltage for the TFT connection push electrode.

[0105] , because the top plate voltage

[0106] This is additional for the voltage supplied to the TFT.

[0048] As illustrated in Figure 2, the active matrix of the propulsion electrode can be configured to be driven by data and gate (select) lines (very similar to the active matrix in a liquid crystal display). The gate (select) lines are scanned for column addressing, while the data lines carry the voltage transferred to the propulsion electrode for electron wetting operations. If no movement is required or if the droplet intends to move away from the propulsion electrode, 0 V will be applied to the (non-target) propulsion electrode. If the droplet intends to move towards the propulsion electrode, an AC voltage will be applied to the (target) propulsion electrode.

[0049] The architecture of the amorphous silicon TFT switching drive electrode is shown in Figure 3. Dielectric

[0308] The dielectric layer must be thin enough and have a dielectric constant compatible with low-voltage AC driving, as is the case with conventional image controllers found in LCD displays. For example, the dielectric layer could comprise a layer of approximately 20-40 nm SiO2 coated with a 200-400 nm plasma-deposited silicon nitride layer. Alternatively, the dielectric could comprise an atomically deposited Al2O3 layer, preferably 20-60 nm thick. TFTs can be constructed by producing alternating layers of different doped a-Si structures and different electrode lines using methods known to those skilled in the art. Hydrophobic layer

[0307] It can be constructed from the materials listed above, such as Teflon® AF and FlurrorPel™, which can be spin-coated onto the dielectric layer.

[0308] Above.

[0050] The circuitry used to connect and / or control the voltages of the top and bottom electrodes can be housed within the top plate itself, the bottom plate (e.g., at the edge of the electrode array), or elsewhere in the device, depending on the requirements and constraints of the current application. As stated above, Cho et al. (Journal of Microelectromechanical Systems, Vol. 12, No. 1, February 2003) provided a physical analysis of how basic droplet manipulation is performed on conventional electronic wetting devices.

[0051] Figure 14 schematically illustrates how a droplet can be cut by selective actuation of an EWoD electrode. During sequential cutting, by activating the electrode on either side and keeping the middle uncharged, the head of the neck is pinched in the longitudinal direction. During pinching, the left and right electrodes are charged, thus reducing their contact angle and increasing the radius of curvature R1. Simultaneously, the electrode at the pinch point is suspended or grounded, maintaining the hydrophobicity of the middle portion. Therefore, the meniscus on the middle electrode begins to contract to maintain the total volume of the neck constant. That is, cutting is initiated by the longitudinal elongation of the droplet and the necking in the middle of the droplet (negative radius of curvature R, also shown in Figure 14). It can be shown that the ratio of the radii of curvature R to R1 follows equation (1): Equation (1) in Vacuum permittivity, γ is the dielectric constant of the dielectric layer, t is the thickness of the dielectric layer, Vd is the applied voltage, d is the height of the gap in the microfluidic region, and γ is the surface tension between the droplet and the filler fluid (see Cho et al.).

[0052] As stated above, the application drive sequence according to this application utilizes a high-density electrode array. Figure 4 is a schematic top view of the reservoir 400 defined by the high-density electrode grid 402. For example, an area with a 1 in² area and a base electrode density resolution of 100 PPI will cover 100 propulsion electrodes. The same area but a higher resolution (e.g., 200 PPI or more) will result in an area with 200 or more propulsion electrodes. It can be seen that the density of the electrode grid allows its pixels to have dimensions smaller than the droplet diameter, such as width, height, or diagonal, which allows for the application of droplets of different sizes and aspect ratios. For example, droplet 404 is equivalent in width and height to a square formed by four electrodes, droplet 406 is larger and equivalent to eight electrodes, and droplet 408 has the same height as droplet 406 but is twice as wide, producing a rectangular shape with an aspect ratio of 2:1. However, embodiments with single-electrode droplets are also covered.

[0053] Figure 5 shows the same reservoir as in Figure 4, with the electrode grid omitted for clarity. The dashed lines indicate the electrode actuation region. It is evident that the difference between Figures 5A and 5B lies in the height of the actuation neck, i.e., the long extension. The reservoir region where electrode actuation occurs is defined as the "frame," which needs to prevent the aqueous fluid from moving uncontrollably away from the reservoir region. A portion of the fluid is driven outside the reservoir to form the actuation "neck," i.e., the extended region ending at the "head," which is the forward-moving edge of the fluid. It is evident that the "pool" is formed on either side of the neck due to excess fluid not contained within the dispensing model. Ideally, the goal is to minimize pool formation while allowing the neck to extend freely and the droplets to separate from the head.

[0054] [Actuation parameters]

[0055] The actuation parameters, including those illustrated in Figure 6, can be used to plan and implement electrode drive sequences for dispensing droplets of the desired size and aspect ratio. The parameter values ​​can be calculated to interpret the shape and other features of the reservoir, droplet, neck, and holder. Each of these features and its associated parameters is then examined.

[0056] Storage container: The storage container is specified to have a certain area (LR·WR) equal to the storage container length (LR) multiplied by its width, wherein the width (WR) of the storage container is parallel to the dispensing direction. The storage container fluid will typically be aqueous and contain surfactants, buffers, proteins such as enzymes, nucleic acid molecules, or other compounds. Dispensing is not limited to aqueous fluids, but other solvents and solutes, such as alcohols, ethers, ketones, aldehydes, etc., can also be dispensed by precisely adjusting the parameters disclosed herein.

[0057] Droplet size: The size of the droplet can be provided as droplet volume or droplet diameter. Alternatively, it can be specified based on the pixel area covered by the droplet on the device surface, for example, by multiplying the area length by its height. In one embodiment, the user can input a specific droplet volume into a programmed device to calculate its corresponding area. In cases where a droplet with the most square possible coverage area is required, its area can be calculated using the following algorithm: (1) Calculate the square root of the volume to obtain the value "X". (2) Rounding floating-point numbers, for example (3) Perform the calculation: X · X, (X+1) · (X-1), X(X+1) (4) The result closest to the initial volume becomes the droplet dimension, for example, by setting LD = X and WD = X+1. (5) Typically, the dimension is smallest in the direction orthogonal to the apex, and is referred to in this paper as head height. [s] 」.

[0058] Neck parameters: except for head height In addition to [s], by means of neck length " [n] defines the neck, which can be set by the user or calculated by the device. The value of [n] should be kept reasonable to avoid exceeding the storage capacity limit. Typically, the product... [n] [·] [s] should not exceed a threshold percentage of the reservoir volume, such as 80% or less. Parameter " [g] [*] 」 Marks the position where the neck begins relative to the length of the gap between the frame edges, and in principle can be zero or negative, so that the neck begins at or even after the frame edge.

[0059] Frame parameters: The frame length should be remembered. [h] is set to the volume of the storage unit. Typically, when the rack extends throughout the entire vertical dimension LR, [h] is approximately 10%-20% of the area occupied by the fluid in the reservoir. Adjustable rack length. [h] to explain changes in reservoir fluid volume and to control the size of the reservoir based on droplet size. In one embodiment, [h] is adjusted proportionally to 1 / D2, where D is the droplet diameter, to tighten the reservoir when dispensing smaller droplets.

[0060] parameter" [g] defines the adjustment spacing used to regulate the decrease in fluid in the reservoir and the placement of the retainer at the location of the residual fluid. For example, if If [g] is always equal to zero, then eventually it will no longer be possible to keep the reservoir fluid in the proper position. Parameter " [h] [*] 」Defines the height of the frame when it differs from LR. This is due to the reduction in overall fluid volume. [h] [ * The value may need to be reduced at the start of the application drive sequence. This will allow fluid to accumulate around the intended location for neck formation. This height [h] [*] This can also be altered when the droplets are pinched and / or broken down, and can be increased beyond its application value according to equation (1). The gap between the frame and the neck. [g] [ * The neck length can be adjusted to handle more viscous or problematic fluids, resulting in fewer restrictive actuations throughout the reservoir. In a non-limiting embodiment, the neck length... [n] is adjusted proportionally to 1 / D to achieve improved delivery of smaller droplets. In another non-limiting embodiment, the head height... [s] is adjusted proportionally to D to achieve the application of droplets of different sizes.

[0061] Size Range and Limitations: Typically, electronically wetted arrays have a grid of square pixels spaced in a regular pattern. However, the method disclosed in this application can be implemented on grid patterns of electrodes and / or pixels based on different geometries, such as triangles, rectangles, or hexagons, and with different dimensions, provided that the spatial and temporal necking as disclosed herein remains feasible. Pixel sizes can be varied for TFT architectures, but there are no fundamental limitations to ensure reservoir operation. Typical pixel values ​​range from 100 micrometers to 1 mm pixel length, but can be extended beyond this range. Similarly, the array can be constructed with variable resolution regions to ensure finer dimensional measurements (e.g., finer resolution regions to induce necking from droplet separation, via, etc.). [s] The parameters [*] are described below.

[0062] The reservoir, rack, neck, and droplet sizes can be specified based on the surface area measured as per the pixel count. The droplet volume should generally not exceed approximately 30% of the reservoir volume, as application may prove problematic with larger volumes. It is preferable not to exceed the array's operating temperature range. Similarly, it is preferable not to exceed the freezing and boiling points of the relevant liquids. Typical ranges for aqueous formulations span from 4°C to 95°C.

[0063] By applying user input to a reference stored in memory, the processing unit can calculate each of the actuation parameters. For example, in the actuation neck... In an embodiment where the length of [n] is adjusted proportionally to 1 / D, the processing unit of the device can apply a reference correlation in the form of equation (2): Equation (2) Where a and b are constants specific to the reference correlation, which may vary depending on the type of fluid used and other characteristics of the current application, such as the measured temperature or surface tension. In some cases, the equation may include terms proportional to other powers of D, such as 1 / D² or D¹ / ², and / or other terms unrelated to other variables specific to the application. Similar considerations are applied to the algorithmic steps to facilitate the calculation of the actuator length and actuator neck height.

[0064] [Generate image and output to electrodes]

[0065] The image corresponding to the storage application event can be generated as a user input implementation and the actuation parameters are calculated in a manner similar to an animation composed of sequential steps. In one embodiment, encoding is assigned to active pixels and inactive pixels. Inactive pixels will ultimately not receive voltage pulses, while active pixels will receive a set of voltage pulses from each output image (referred to herein as a "waveform"). The image is then transferred to the controller in waveform form, specifying the voltage pulses to be applied to the active pixels.

[0066] In an active matrix device, the controller uses active matrix scanning to drive pixels to their individual voltages. Each image corresponds to a separate step in the memory delivery routine. Routing can continue for multiple steps / images up to droplet delivery. Each image is implemented by multiple voltage pulses or "frames," where active pixels are driven to a set voltage, while non-active pixels typically remain at 0 V. Voltage pulses can span a given positive or negative range, typically within ±30 V or ±40 V on a TFT array. As illustrated in Figure 15A, the drive sequence can include both positive and negative voltage pulses. The voltage pulse frequency is defined by the duration for which the active pixel receives voltage pulses of specific voltage and polarity.

[0067] [Example]

[0068] Figure 7 illustrates an exemplary droplet application method.

[0700] This allows for the calculation and implementation of specific top and bottom plate electrode driving sequences based on the diameter and aspect ratio of the droplets to be dispensed in the microfluidic system. In the steps...

[0702] In this process, the user inputs the desired droplet diameter and aspect ratio in the form of instructions stored in a computer-readable medium accessible by the device's processing unit. The user can also input other relevant variables that affect the actuation parameters, such as the viscosity and surface tension of the aqueous fluid in the droplet.

[0069] The instructions cause the processing unit to execute the algorithm stored in the computer-readable medium and calculate actuation parameters specific to the characteristics of the desired droplet, including neck and frame parameters, such as the width of the frame, the length of the neck, and the height of the head.

[0704] Under the control of the processing unit or by the user before or during the dispensing process, the parameters can be calculated as a function of the input variables based on one or more references that can be saved to memory locations.

[0070] The processing unit then generates an image corresponding to the application.

[0706] And calculate the polarity, frequency, and amplitude of each pulse in the corresponding waveform.

[0707] Subsequently, the processing unit outputs a waveform to the controller.

[0708] ), and the controller outputs a signal to the driver of the propulsion electrode (

[0710] In cases where the substrate includes a TFT electrode array, the controller outputs a gate line signal to the gate line driver and a data line signal to the data line driver, thereby driving the desired propulsion electrode. The selected propulsion electrode is then driven to perform a droplet application drive sequence.

[0712] ).

[0071] Figure 8 shows the process starting from configuration. [A] is a schematic diagram of an exemplary distribution model, in which fluid is collected vertically toward the center. The configuration is selected as appropriate. [B] [ * In the configuration, fluid moves to the front end of the reservoir and is in the configuration. [B] In this configuration, the framework and neck are formed. Subsequently, in the configuration... In [C], droplet decomposition begins at the head. The appropriate configuration is selected based on the situation. [D] [ * In the configuration, the droplet requires an additional step to move away from the head before pulling the neck back to the reservoir; this is referred to in this paper as the "timing neck" stage. Finally, in the configuration... In [D], the reservoir reforms and the droplets travel further.

[0072] Figures 9-13 illustrate individual stages of the dispensing model in Figure 8. Stage 1, illustrated in Figure 9, involves several operations of gathering fluid in a reservoir. This can be achieved by vertically gathering it (A), and subsequently collecting any liquid from the back (B) and moving it to the front (C). Typically, the liquid at the front of the designated reservoir area is the preferred starting point for the dispensing operation. The size of the gathering pattern (shown in magenta) typically extends at least one full length or width of the reservoir area, with other dimensions adjusting with the residual volume of the reservoir, and in the cases of B and C, being large enough to extend at least 20% beyond the liquid edge. For vertical gathering (or a direction orthogonal to dispensing), the gathering pattern covers approximately 50% of the reservoir's length (horizontally) and vertical space. It should be noted that reservoirs can be positioned for both vertical and horizontal dispensing, and therefore these definitions may vary depending on orientation.

[0073] Figure 10 illustrates stage 2, in which the frame and neck are generated, followed by neck stretching. As disclosed above, several actuation parameters are related to the frame and neck. The neck begins to shorten (approximately the size of the target droplet) and then extends outward in the dispensing direction until it reaches the specified neck length. The neck converges around the vertical direction, and as stated above, the parameters... [g] [ *The value can be set so that the neck begins precisely at the edge of the holder. Typically, the neck extends in the application direction by a distance equal to approximately half the desired droplet diameter. However, this value can be as small as that of a single pixel electrode.

[0074] Figure 11 illustrates stage 3, where droplet decomposition begins once the neck is fully extended. The designated deactivation area, showing in red, separates the reservoir liquid from the desired droplet. To initiate decomposition, the area is deactivated by suspending or grounding the electrode within it (A), and the droplet is continuously moved to the right in a minimum step size typically one pixel, or half the pixel size in the application direction (B). The final step involves actuating the area equivalent to completely traversing the neck and any remaining fluid in the direction orthogonal to the application direction to retract the reservoir. Simultaneously, the droplet moves further away from the reservoir (C).

[0075] In the variation of stage 3 illustrated in Figure 12A, step B adds multiple steps and moves the droplet further before pulling it back into the reservoir, thereby creating an extended "timing neck." Through this strategy, the negative radius of curvature R increases to R*, which helps to break up the droplet (Figure 12B). Parameter " [t] defines the number of additional steps that can be used for droplet delivery before pulling the neck back into the reservoir.

[0076] In another variation of Stage 3, as illustrated in Figure 13A, the ability of the necking portion obtained through high-density electrodes in two dimensions can be used to achieve improved control over the droplet fragmentation step. Specifically, the head height... [s], that is, the dimension of the forward-moving neck, which is orthogonal to the forward-moving direction, can be increased to a new "forward-moving decomposition height" greater than the original by activating electrodes on either side of the neck. [s] [ * As shown in equation (1), in order to split the neck, R should gradually become negative, thus requiring a larger R1 (by increasing...). [s] to [s] [ * (, ] to obtain) in order to achieve a more efficient decomposition (Figure 13B). Parameter " [s] [*] can be referred to as the new height of the neck side orthogonal to the direction of application. [s] [ * , ] greater than The length of [s] can be specified based on the pixel electrode or be the original head height. [s] in percentage form.

[0077] Those skilled in the art will readily recognize that numerous changes and modifications can be made to the specific embodiments of the invention described above without departing from the scope of the invention. Therefore, the entire foregoing description is to be interpreted as illustrative rather than restrictive.

[0078] All contents of the aforementioned patents and applications are incorporated herein by reference in their entirety. In the event of any inconsistency between the contents of this application and any of the patents and applications incorporated herein by reference, the contents of this application shall prevail to the extent necessary to resolve such inconsistency.

[0079] 100:EWoD 102: Filler fluid 104: At least one aqueous droplet 105: Propulsion Electrode 106: Top electrode 107: Hydrophobic coating 108: Dielectric layer 307: Hydrophobic layer 308: Dielectric layer 400: Storage 402: High-density electrode grid 404: Droplets 406: Droplet 408: Droplet 700: Exemplary Droplet Application Method 702: Steps 704: Steps 706: Steps 707: Steps 708: Steps 710: Steps 712: Steps g: Adjust the spacing g*: Gap length h: Frame length h*: Frame height n: Neck length R: Radius of curvature R*: Radius of curvature R1: Radius of curvature s: head height s*: Move the decomposition height forward

Claims

1. A method for applying droplets to a digital microfluidic system, the system comprising: (a) a substrate including: a bottom electrode array including a plurality of digital microfluidic propulsion electrodes; and a first dielectric layer covering the bottom electrode array; (b) a top plate including: a common top electrode; and a second dielectric layer covering the common top electrode; (c) a processing unit operably programmable for performing a microfluidic actuation method; and (d) a controller operably coupled to the processing unit, the common top electrode, and the bottom electrode array, wherein the controller is configured to provide a propulsion voltage between the common top electrode and the substrate propulsion electrodes; the microfluidic actuation method comprising: receiving input commands in the processing unit, the input commands relating to droplet diameter and aspect ratio; and calculating in the processing unit actuation parameters including: length of actuator frame, length of actuator neck, and height of actuator head, for applying droplets having the diameter and aspect ratio of the input commands; Electrode actuation commands are output from the processing unit to the controller, the electrode actuation commands relating to an application drive sequence for implementing calculation of actuation parameters; the application drive sequence is executed on the propulsion electrodes to: shape fluid in a reservoir to form an actuation frame and an actuation neck; cause the droplet to disintegrate from the head of the actuation neck; and cause the fluid in the actuation neck to return to the reservoir, wherein the electrodes have a dimension smaller than the diameter of the droplet.

2. The method of dispensing droplets as claimed in claim 1, wherein the length of the actuator is calculated based on an equation that responds to at least the input droplet diameter and associates the droplet diameter with the length of the actuator.

3. The method of dispensing droplets as claimed in claim 1, wherein the length of the actuating neck is calculated according to an equation that responds to at least the diameter of the input droplet and associates the droplet diameter with the length of the actuating neck.

4. The method of dispensing droplets as claimed in claim 1, wherein the height of the actuating head is calculated according to an equation that responds to at least the diameter of the input droplet and associates the droplet diameter with the height of the actuating neck.

5. The method of dispensing droplets as claimed in claim 1, wherein the actuation parameters further include one or more of the following: reservoir height, adjustment space of the frame, length of the actuation frame, height of the actuation neck, frame spacing, amount of residual fluid in the reservoir, and length of the gap between the actuation frame and the actuation neck.

6. The method of dispensing a droplet as claimed in claim 1, further comprising forming a timing neck to provide the droplet with additional time to move away from the actuation neck.

7. The method of dispensing droplets as claimed in claim 1, further comprising raising the height of the actuating head to a forward dissolution height before dissolving the droplets from the head of the actuating neck.

8. The method of dispensing droplets as claimed in claim 1, further comprising reducing the height of the holder to cause the fluid to accumulate around the location where the actuation neck is formed.

9. A digital microfluidic system comprising: (a) a substrate including: a bottom electrode array including a plurality of digital microfluidic propulsion electrodes; and a first dielectric layer covering the bottom electrode array; (b) a top plate including: a common top electrode; and a second dielectric layer covering the common top electrode; (c) a processing unit; and (d) a controller operatively coupled to the processing unit, the common top electrode, and the bottom electrode array, wherein the controller is configured to provide a propulsion voltage between the common top electrode and the substrate propulsion electrodes; and wherein the processing unit is operatively programmed to: receive input commands relating to droplet diameter and aspect ratio; and calculate actuation parameters including: the length of an actuator frame, the length of an actuator neck, and the height of an actuator head, for applicating droplets having the diameter and aspect ratio of the input commands; The output electrodes actuate the controller, and the electrode actuation commands relate to a dispensing drive sequence for implementing the calculated actuation parameters to dispense a droplet having the diameter and aspect ratio of the input commands; wherein the electrodes have a dimension smaller than the droplet diameter.

10. The digital microfluidic system of claim 9, wherein the processing unit is operably programmed to calculate the length of the actuator according to an equation that responds to at least the input droplet diameter and correlates the droplet diameter with the length of the actuator.

11. The digital microfluidic system of claim 9, wherein the processing unit is operably programmed to calculate the length of the actuation neck according to an equation that responds to at least the input droplet diameter and correlates the droplet diameter with the length of the actuation neck.

12. The digital microfluidic system of claim 9, wherein the processing unit is operably programmed to calculate the height of the actuator head using an equation that responds to at least the input droplet diameter and correlates the droplet diameter with the height of the actuator head.

13. The digital microfluidic system of claim 9, wherein the actuation parameters further include one or more of the following: reservoir height, adjustment space of the frame, length of the actuation frame, height of the actuation neck, frame spacing, amount of residual fluid in the reservoir, and gap length between the actuation frame and the actuation neck.

14. The digital microfluidic system of claim 9, wherein the processing unit is further operably programmable to form a timing neck to provide the droplet with additional time to move away from the neck.

15. The digital microfluidic system of claim 9, wherein the processing unit is further operablely programmable to raise the height of the actuator head to a forward disintegration height before disintegrating the droplet from the head of the neck.

16. The digital microfluidic system of claim 9, wherein the processing unit is further operablely programmable to reduce the height of the frame so that the fluid accumulates around the location formed around the neck.

17. The digital microfluidic system of claim 9, wherein the base plate further includes a transistor active matrix backplane, wherein each transistor of the backplane is operatively connected to a gate driver, a data line driver and a drive electrode.

18. The digital microfluidic device of claim 17, wherein the transistor of the backplane is a thin-film transistor (TFT).

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