Apparatus, system and method for droplet manipulation

Abstract

The invention relates to an apparatus for droplet manipulation, comprising: a driving module configured to receive a control signal and operate an on-medium electrowetting chip configured to manipulate droplets; a droplet detection system comprising a light source and a lensless imaging device configured to obtain at least one image comprising a droplet in the electro-wetting-on-medium chip; and a control unit configured to receive the at least one image from the droplet detection system and send a control signal to the driving module. The control unit is further configured to adapt the at least one control signal in accordance with the at least one image. The invention also relates to a system comprising said instrument and an electrowetting-on-dielectric chip configured to receive signal electricity from a driver module.

Description

Technical Field

[0001] The present invention generally relates to an instrument, system and method for droplet manipulation, and more particularly to an instrument, system and method for droplet manipulation by electrowetting-on-dielectric (EWOD). Background Art

[0002] Droplet manipulation has crucial commercial and scientific potential for many biomedical applications. The current state of droplet manipulation is limited in terms of throughput, cost, and functionality.

[0003] D. Cyril et al. explain a known technique for tracking droplets on an EWOD chip in the document WO2016 / 174523A1. Summary of the Invention

[0004] The present invention is set forth in the appended claims.

[0005] The object of the present invention is to overcome at least in part the limitations of the prior art. Specifically, the object of the present invention is to provide an instrument, system and method for effective droplet manipulation.

[0006] In a first aspect, the present invention relates to an instrument for droplet manipulation, comprising: a driving module configured to receive a control signal and operate an electrowetting-on-dielectric (EWOD) chip, wherein the EWOD chip is configured to manipulate droplets; a droplet detection system comprising a light source and a lensless imaging (LFI) device configured to obtain at least one image comprising droplets in the EWOD chip; and a control unit configured to receive at least one image from the droplet detection system and to send a control signal to the driving module; wherein the control unit is further configured to adapt at least one control signal based on the at least one image. The instrument of the first aspect has several advantages. First, the lensless imaging device can capture microscopic images with a large field of view (FOV), and thus can continuously and automatically monitor and manipulate multiple droplets in parallel. Second, the images captured by the lensless imaging device enable adjustment and adaptation of one or more subsequent control signals to manipulate the droplets. Third, the instrument is compatible with various customized EWOD chips.

[0007] Any feature of the first aspect may be as described correspondingly in the second aspect.

[0008] In a second aspect, the present invention relates to a system that includes the instrument and also includes an electrowetting-on-dielectric chip. The system is preferably in a replaceable cartridge and is configured to receive signals from a drive module. The system has the advantage of being compatible with various replaceable cartridges. According to an exemplary embodiment, the replaceable cartridge is a disposable cartridge. According to an exemplary embodiment, the replaceable cartridge is a reusable cartridge.

[0009] According to an example embodiment, at least one of the adapted control signals is for instructing the drive module to move a droplet to a predetermined position. The advantage of this exemplary embodiment is that the control unit automatically determines the path for moving the droplet to the predetermined position based on the image captured by the lensless imaging device.

[0010] According to an example embodiment, the control unit is configured to extract the position of a droplet from at least one image, and at least one control signal is adapted based on the position. This has the advantage of allowing the positions of multiple droplets to be captured in the image and the EWOD pixels to be captured in the absence of a droplet therein.

[0011] According to an example embodiment, the control unit is configured to adapt the control signal such that if the movement of the droplet after the first control signal is different from a predetermined first path, at least one subsequent control signal is sent to move the droplet along a second path different from the first path to the predetermined position. The advantage of this example embodiment is that during the movement of the droplet, the path for moving the droplet to the predetermined position can be adapted and changed to a different path.

[0012] According to an example embodiment, the at least one image includes a holographic image. The advantage of this example embodiment is that the control signal can be adapted based on the raw data (i.e., hologram) from the lensless imaging device. This allows the control unit to quickly adapt the control signal based on the hologram captured by the lensless imaging device because the processing speed on the hologram is not constrained by the computing core.

[0013] According to an example embodiment, the at least one image includes a two-dimensional (2D) or three-dimensional (3D) image. According to an example embodiment, the 2D or 3D image can be a reconstructed image from the raw data, i.e., a hologram. According to an example embodiment, the 2D or 3D image can be captured by a different imaging device. Advantageously, the 2D or 3D image includes more information than the hologram. Therefore, the control signal can be adapted according to the reconstructed image.

[0014] According to an example embodiment, the control unit is configured to adapt at least one control signal based on at least one variable obtained by the control unit from at least one image, each of the at least one variable representing a different property selected from one or more of the spatial distributions of size, shape, spectral absorptivity, spectral transmittance, turbidity, viscosity, and at least one of the foregoing properties of the droplet. It is advantageous for the lensless imaging device to capture properties including droplet morphology in at least one image. The properties captured in the image can be further used as variables for adapting the control signal.

[0015] According to an example embodiment, at least one control signal is adapted to instruct the drive module to mix at least two droplets into one droplet.

[0016] According to an example embodiment, the control unit is configured to adapt at least one control signal to continue or stop the mixing of droplets based on at least one variable obtained by the control unit from at least one image, the at least one variable representing the spatial distribution of at least one property or combination of properties selected from one or more of the size, shape, spectral absorptivity, spectral transmittance, turbidity, and viscosity of the droplet. According to an example embodiment, the image is a 3D image. An advantage is that the one or more images can be used to mix or combine droplets to automatically achieve homogeneity.

[0017] According to an example embodiment, the control unit is configured to adapt at least one control signal based on at least one variable obtained by the control unit from the at least one image, each of the at least one variable representing an object in the droplet. Advantageously, features in the droplet image, i.e., objects, are captured by the lensless imaging device in at least one image. The features in the image can be further used as variables for adapting the control signal.

[0018] According to an example embodiment, the object is a cell. An advantage is that particles such as cells in the droplet can be captured in the image. Such particles in the image can be further used as variables for adapting the control signal.

[0019] According to an example embodiment, the object is a bubble. An advantage is that defects such as bubbles can be captured in the image. Such defects in the image can be further used as variables for adapting the control signal.

[0020] According to an example embodiment, the object is a different immiscible droplet, i.e., another droplet that is insoluble in the droplet. An advantage is that the properties of the droplet and the different droplet therein can be extracted. Advantageously, any differences between at least two droplets can be captured by the lensless imaging device in at least one image. Such differences in the image can be further used as variables for adapting the control signal.

[0021] According to an exemplary embodiment, the control unit is configured to adapt at least one control signal based on at least one variable obtained by the control unit from at least one image, each of the at least one variables representing a different defect in the electrowetting-on-dielectric chip. An advantage of this exemplary embodiment is that the image also captures the defects on the EWOD chip. These defects indicate the fault locations on the EWOD chip. Thus, when manipulating the droplets, the control signal can be adapted to avoid such locations.

[0022] According to an exemplary embodiment, the electrowetting-on-dielectric chip includes a plurality of electrowetting-on-dielectric layers, wherein the control unit is configured to adapt at least one control signal based on at least one variable obtained by the control unit from a plurality of holographic images by using focus adjustment, each of the at least one variables representing whether a droplet is present in a different electrowetting-on-dielectric layer. An advantage is that droplets can be manipulated simultaneously and in parallel on multiple stacked EWOD layers. This allows for more efficient and faster droplet manipulation.

[0023] Any feature of the second aspect may be as described correspondingly in the first aspect.

[0024] In a third aspect, the present invention relates to a method for droplet manipulation in an instrument according to an embodiment of the specification, comprising the steps of: a. causing a control unit (104) to receive at least one image including a plurality of droplets; b. causing the control unit (104) to generate control signals for simultaneously manipulating the droplets by electrowetting based on the received at least one image. It is advantageous to manipulate a plurality of droplets simultaneously using this method. According to an exemplary embodiment, at least 1000 droplets are simultaneously manipulated using this method. According to an exemplary embodiment, at least 10,000 droplets are simultaneously manipulated using this method.

[0025] Any feature of the third aspect may be as described correspondingly in the first or second aspect.

[0026] In a fourth aspect, the present invention relates to a computer program comprising instructions for causing an instrument according to the first aspect to perform the steps of the method according to the third aspect.

[0027] In a fifth aspect, the present invention relates to a computer-readable medium having stored thereon the computer program according to the fourth aspect. Description of the Drawings

[0028] Figure 1 A block diagram of a first example is shown, which shows an instrument for droplet manipulation in combination with a top view of an EWOD chip.

[0029] Figure 2 A block diagram of a second example of a system for droplet manipulation is shown, as well as a side view of an EWOD chip.

[0030] Figure 3 A block diagram showing a third example of a system for droplet manipulation, and illustrations of droplet images in a hologram and a reconstructed image.

[0031] Figure 4 A block diagram showing a fourth example of a system for droplet manipulation, and illustrations of images of the extracted properties of the droplet.

[0032] Figure 5 A block diagram showing a fifth example of a system for droplet manipulation, and illustrations of images of the extracted properties of the droplet captured at time intervals.

[0033] Figure 6 A block diagram showing a sixth example of a system for droplet manipulation, and illustrations of images of droplets with an object therein.

[0034] Figure 7 A block diagram showing a seventh example of a system for droplet manipulation, and illustrations of images of droplets with a bubble as a defect.

[0035] Figure 8 A block diagram showing an eighth example of a system for droplet manipulation, and a side view of an EWOD chip having two EWOD layers.

[0036] Figure 9 An enlarged view of a droplet detection system and an EWOD chip showing a ninth example of a system for droplet manipulation.

[0037] Figure 10 An enlarged view of a solution loaded on an EWOD chip.

[0038] Figure 11 An enlarged view showing a droplet generation process. Detailed Description

[0039] The present disclosure will be further clarified by the following description and the accompanying drawings. Various exemplary embodiments are described herein with reference to the following drawings, in which the same numbers represent the same entities. The described drawings are schematic and not restrictive. Further, any reference numerals in the claims should not be construed as limiting the scope of the present disclosure. Moreover, the same reference numerals refer to the same or similar elements in different drawings.

[0040] The term "comprising" as used in the claims should not be construed as being limited to the means listed thereafter; it does not exclude other elements or steps. Thus, the term should be construed as specifying the presence of the stated features, integers, steps or components as mentioned, but not excluding the presence or addition of one or more other features, integers, steps or components, or groups thereof. The term "comprising" thus encompasses the case where only the stated features are present (and can therefore always be replaced by "consisting of" to limit the scope to the stated features) as well as the presence of these features and one or more other features. The word "comprising" according to the present invention thus also includes, as an embodiment, the absence of other components. Thus, the scope of the expression "an apparatus comprising means A and B" should not be construed as being limited to an apparatus consisting only of components A and B. This means that for the purposes of the present invention, the relevant components in the apparatus are only A and B.

[0041] The term "a" should be construed as a functional word before a mass noun to denote a particular type or instance. It should not be construed as a dummy word before a singular noun referring to one object.

[0042] The term "instrument" refers to a tool or device designed for droplet manipulation.

[0043] A droplet refers to a volume of a first fluid in the range of 30 picoliters to 100 microliters in a second fluid, where the first and second fluids are immiscible. According to an example embodiment, the first fluid is aqueous and the second fluid is oil-based, such as silicone oil. According to an example embodiment, the droplet may also include particles and / or objects, such as biological cells, bubbles, another droplet, etc. Droplet manipulation generally refers to actions applied to the droplet, including but not limited to: changing the position of the droplet, interacting with the droplet, and causing a change in the properties of the droplet.

[0044] As Figure 1 shown, the present invention relates to an instrument (100) comprising a drive module (101), a droplet detection system (103) and a control unit (104). The instrument (100) is compatible for use with an EWOD chip (102).

[0045] As Figure 2The exemplary EWOD chip shown includes an EWOD layer (25), a transparent cover (24), and a thin film transistor (TFT) backplane (26). A distance is left between the EWOD layer (25) and the transparent cover (24) to form an operating space in which to manipulate droplets and place fluids around the droplets. This operating space may be referred to as the electrowetting layer. The operating space typically has a height of 10 to 1000 μm to allow droplet manipulation. According to one example embodiment, the transparent cover (24) is a glass cover. According to one exemplary embodiment, the EWOD chip further includes jetting technology, such as reservoirs for providing droplets and fluids. According to one exemplary embodiment, the reservoir contains biochemical materials, such as cell culture media, buffers, oils, water, etc. According to one exemplary embodiment, the biochemical materials can be supplied to the reservoir by an attached fluid pump or can be supplied manually or automatically by a pipette. Such biochemical materials can be contained, for example, in tubes, containers, plates, syringes attached to the fluid pump.

[0046] The EWOD chip further includes an electronic interface (21) configured to receive control signals from a drive module (101). The EWOD layer (25) includes an electrode array. The TFT backplane includes electronic connections between each electrode in the drive module (101) and the electrode array. The TFT backplane has electrode pins to make an electrical connection with the instrument. Droplets are manipulated by the electrodes such that the wetting properties of the droplets are changed and the contact angle of the droplets on the surface of the EWOD chip is changed due to electrostatic effects controlled by the electrode array. The EWOD chip may further include one or more carriers (22) for support purposes.

[0047] According to one example embodiment, the drive module (101) includes an electronic interface compatible with the corresponding electronic interface on the EWOD chip. Such a drive module (101) receives control signals from a control unit. According to one example embodiment, the drive module (101) is a general-purpose computer, an ASIC chip, or an FPGA module.

[0048] The instrument (100) includes a droplet detection system (103). The droplet detection system (103) includes a lensless imaging device (31). The term "lensless imaging device" may be interchangeable with "lensless holographic imaging device". According to an example embodiment, the droplet detection system (103) further includes one or more other imaging devices, such as a CCD camera and / or a fluorescence detector. The droplet detection system includes a light source (32) for emitting light onto an illumination area on the EWOD chip. According to an example embodiment, the light source is configured to irradiate at least a portion of the top surface of the EWOD chip. The top surface generally refers to the surface of the EWOD chip that receives light from the light source. According to an example embodiment, the top surface is a transparent cover. According to another example embodiment, the top surface is a TFT backplane. Since the EWOD chip is substantially transparent to light, the top surface or the bottom surface may be interchangeable in context. The EWOD chip is placed at an intermediate point along the optical path between the light source and the lensless imaging device (31). The glass cover (24), the operation space, the EWOD layer (25), and the TFT backplane (26) are substantially transparent, such that the light emitted from the light source can be detected by the lensless imaging device (31). The lensless imaging device (31) includes an imager chip (312) and an imager PCB (311). According to an example embodiment, the temporal resolution of the lensless imaging device may be less than or equal to 1 millisecond.

[0049] An exemplary lensless imaging device (31) has a field of view (FOV) of 20 mm 2 . The illumination area is greater than or equal to the FOV of the lensless imaging device (31). According to an example embodiment, the droplet detection system (103) includes a plurality of lensless imaging devices.

[0050] The lensless imaging device (31) captures at least one raw hologram of at least a portion of the illumination area. A holographic image of the entire FOV is captured. It includes a diffraction pattern on the FOV and a corresponding diffraction pattern of the droplet of interest in the EWOD chip (102). The holographic image may be further reconstructed into a two-dimensional (2D) and / or three-dimensional (3D) image. According to an example embodiment, the droplet detection system includes a processing unit configured to reconstruct the holographic image into a 2D and / or 3D image.

[0051] The control unit (104) receives at least one image from the droplet detection system. According to an example embodiment, the image is a holographic image. According to an example embodiment, the image is a reconstructed 2D or 3D image. According to an example embodiment, the control unit (104) includes a central processing unit for processing the image received from the droplet detection system. According to an example embodiment, the central processing unit is configured to reconstruct the holographic image into a 2D and / or 3D image. According to an example embodiment, the central processing unit is in a general-purpose computer.

[0052] The control unit (104) sends a control signal to the drive module (101) to command the electrodes in the EWOD chip. The control signal commands the electrodes to change the contact angle of the droplet on the EWOD layer, so that the droplet is controlled by the electrostatic effect conducted by the electrodes. The control signal can be, but is not limited to: transferring droplets, splitting droplets, dispensing droplets, merging droplets,....

[0053] At least one of these control signals is adapted according to the at least one image.

[0054] The features captured in the image are used as variables in the function for adapting the control signal. According to an example embodiment, the feature is the position of the droplet captured in the image. According to an example embodiment, such a feature can be at least one parameter of the interrelationship of at least two droplets captured in the image, such as distance. According to an example embodiment, such a feature can be at least one parameter of the properties of the droplet corresponding to the diffraction pattern of the droplet of interest extracted from the received holographic image. According to an example embodiment, multiple features are considered as variables for adapting the control signal.

[0055] According to an example embodiment, the control unit (104) is configured to extract the position of the droplet from the at least one image, and at least one control signal is adapted based on this position.

[0056] The electrodes may be damaged during manufacturing or operation. The damage can be partial breakdown of the hydrophobic coating or dielectric layer. It can also be complete dielectric breakdown. According to an example embodiment, during operation, the control unit (104) sends a control signal to command the droplet to be transferred along a first path to a predetermined position. When the droplet movement is different from the predetermined path defined by the control unit, a faulty electrode is detected. The control unit (104) will design a second path to the predetermined position, the second path being different from the first path, and adapt subsequent control signals to guide the droplet along the second path. So that the droplet is still delivered to the predetermined position, but the path via the faulty electrode is avoided. A faulty electrode refers to an electrode that cannot properly change the wetting property of the droplet of interest to move it on the electrode. The failure of the electrode may be caused by damage to the electrode itself or the circuit connected to the electrode.

[0057] According to an example embodiment, during operation, the control unit (104) sends a control signal to command the droplet to be transferred along a first path to a predetermined position, but the last electrode, i.e., the predetermined position, fails. The control unit (104) will dispense the droplet to a second position different from the predetermined position. According to an example embodiment, the predetermined position is in a predetermined area, and the second position can be a different position in the same predetermined area that has not been assigned to another droplet.

[0058] According to an example embodiment, the control unit (104) is configured to perform an EWOD inspection on the defects. The control unit (104) will adapt at least one control signal according to at least one variable obtained by the control unit (104) from at least one image, and each of the at least one variables represents a different defect in the electrowetting-on-dielectric chip (102) on the medium. According to an example embodiment, the defect is damage to the dielectric layer. For example, when dielectric breakdown causes small holes in the device, the damaged area including the electrode is visible in the captured image, such as the edge of the missing part of the dielectric layer or the different gray levels of the missing part of the dielectric layer compared to the surrounding undamaged layer. In another example, when a small area of the hydrophobic layer or dielectric layer is peeled off, the resulting thickness change can be visualized in the captured image, showing a different local light density compared to the functioning electrode. The difference in light density of the electrode or its surroundings can be a variable for adapting at least one subsequent control signal. According to an exemplary embodiment, such an EWOD inspection can be performed before any droplet is loaded onto the EWOD chip. In this way, all control signals are adapted to avoid using damaged areas and faulty electrodes during operation. According to an exemplary embodiment, such an EWOD inspection can be performed during operation, so that any breakdown of the electrode can be observed in a timely manner. The control signal is adapted to the defect detected in the EWOD inspection during operation.

[0059] As Figure 3 shown, the light source (32) in the droplet detection system (103) emits light towards the illumination area on the EWOD chip (102). The lensless imaging device (32) captures a holographic image from the interference of light. The control unit (104) receives the holographic image and / or the reconstructed 2D / 3D image from the droplet detection system (103). At least one control signal is adapted according to the at least one image. The control signal is sent to the EWOD chip (102). Example diagrams of the holographic image and the reconstructed 2D image of the droplet of interest are given.

[0060] According to an example embodiment, the image is a holographic image. Features captured in the original holographic image, such as the position, size, shape, refractive index, turbidity, etc. of the droplet. These features can be discovered by a supervised model or an analytical model developed on the hologram. Can be directly used as variables for adapting the control signal. This allows for the rapid adaptation of the control signal because the processing speed is not constrained by the computing cores required for the reconstruction process.

[0061] According to an example embodiment, the image is a reconstructed image from a holographic image. The reconstruction process requires numerical calculations on the two-dimensional image obtained from the image sensor. Such numerical calculations include, for example, deconvolution, wave or beam propagation, transformation (such as Fourier transform), two-dimensional filtering operations (such as denoising), etc. The reconstructed image reveals the physical characteristics of the droplets.

[0062] According to an example embodiment, the control unit (104) receives the holographic image and the reconstructed image.

[0063] According to an example embodiment, the image can be processed as a whole. According to an example embodiment, the image can be divided into sub-regions, and the sub-regions are processed separately. Thus, the processing can be performed in parallel for different sub-regions.

[0064] The instrument (100) is compatible for use with an EWOD chip. According to an example embodiment, the EWOD chip is in a replaceable cartridge. The cartridge is reusable or disposable. When the cartridge is a disposable device, the instrument (100) can be used with multiple cartridges one after another for multiple experiments. When the cartridge is reusable, the cartridge can optionally be refilled between experiments. The refilling step ensures that the surface properties of the EWOD chip are restored. According to an exemplary embodiment, the instrument (100) provides a mechanical slot in which the user can mount the cartridge onto the instrument. The EWOD chip is configured to receive signals from the drive module (101) of the instrument (100). The EWOD chip includes a corresponding electronic interface to communicate with the drive module (101).

[0065] According to an example embodiment, at least one adapted control signal is for instructing the drive module (101) to move the droplet to a predetermined position. The detection of a feature or a change in a feature can be used as a trigger event to send the droplet to a predetermined position and group the droplets in a predetermined area.

[0066] As Figure 4 shown, the light source (32) in the droplet detection system (103) emits light towards the illumination area on the EWOD chip (102). The lensless imaging device (32) captures the holographic image from the interference of the light. The control unit (104) receives the holographic image and / or the reconstructed 2D / 3D image from the droplet detection system (103). At least one control signal is adapted according to the at least one image. According to an example embodiment, the entire image containing multiple droplets can be processed to adapt subsequent control signals. According to an example embodiment, the original holographic image can be divided into predefined sub-regions in which droplets are present. These sub-regions can be processed independently. The control unit (104) can send control signals to the sub-regions.

[0067] The control signal is sent to the EWOD chip (102). Example diagrams of the properties of the droplets of interest are given, such as size, shape, spectral transmittance, spectral absorptance, and turbidity, as well as the spatial distribution of some of these properties within the droplet. Any one or a combination of these characteristics can be used as a variable for adapting the control signal.

[0068] According to an example embodiment, the control unit (104) is configured to adapt at least one control signal based on at least one variable obtained by the control unit (104) from the at least one image, each of the at least one variable representing a different property selected from the size, shape, spectral absorptance, spectral transmittance, turbidity, and viscosity of the droplet.

[0069] According to an example embodiment, the at least one adapted control signal is adapted to instruct the drive module (101) to mix at least two droplets into one droplet.

[0070] According to an example embodiment, the control unit (104) is configured to adapt at least one control signal to continue or stop mixing droplets based on at least one variable obtained by the control unit (104) from the at least one image, the at least one variable representing the spatial distribution of at least one property or combination of properties selected from one or more of the size, shape, spectral absorptance, spectral transmittance, turbidity, and viscosity of the droplet. According to an example embodiment, "droplet mixing" refers to the process of droplet coalescence by electro-wetting to mix droplets. In some cases, "droplet mixing" may be interchangeable with "droplet coalescence".

[0071] According to an example embodiment, the size of the droplet of interest can be extracted from the image. According to an exemplary embodiment, a predetermined value (optionally with a margin value) can be stored in the control unit and compared with the measured size of the droplet. If the size of the droplet does not meet the requirements, the droplet can be moved to a predetermined position for further study.

[0072] According to an example embodiment, the shape of the droplet of interest can be extracted from the image. According to an example embodiment, the volume of the droplet can be estimated. According to an example embodiment, the surface shape of the droplet can be estimated from the diffraction pattern captured in the 3D image of the droplet. The estimated surface shape provides an indication of the surface tension of the droplet and thus the wetting characteristics can be estimated from the 3D image. Any one or a combination of these characteristics can be used as a variable for adapting the control signal.

[0073] According to an example embodiment, the spectral absorptivity or transmittance of a droplet can be revealed from an image. When the droplet scatters and / or absorbs light to be detected by a lensless imaging device (31), the diffraction effect can be quantified by reconstructing the fringe pattern of the hologram. Accordingly, the attenuation coefficient of the droplet solution can be estimated and used as a variable for adapting a control signal.

[0074] The absorption of the droplet solution can be spectrally related. At different wavelengths within the visible and NIR spectral ranges, the absorption properties of light may be different (image sensor sensitivity). According to an example embodiment, a light source (32) can apply multiple different wavelengths, and the lensless imaging device (31) captures holographic images at different wavelengths. The reconstructed images of the droplet at different wavelengths reveal the spectral absorption of the droplet. According to an example embodiment, the dilution of the substance within the droplet is a characteristic. According to an exemplary embodiment, the presence or absence of a certain soluble substance having the chemical property of absorbing light within a specific spectral window width ranging from 1 nm to 500 nm can be estimated as a characteristic. According to an exemplary embodiment, the change in absorption during a predetermined time period can be observed as a characteristic to monitor chemical or biological reactions inside the droplet. Any one of these characteristics or a combination thereof can be used as a variable for adapting a control signal.

[0075] According to an example embodiment, the light source (32) includes a plurality of sub-light sources that illuminate the droplet at unique and different wavelengths. According to an example embodiment, a single light source is used for illumination at multiple and different wavelengths.

[0076] According to an example embodiment, the turbidity property can be evaluated from the image. The turbidity and blurring caused by the first fluid in the droplet can be detected because the light from the droplet will be scattered, and this will result in a diffraction pattern. The degree of turbidity can be associated with the optical feature holographic image or the reconstructed image of the holographic image. The degree of turbidity can be defined, for example, in formazin turbidity units (FNU) or nephelometric turbidity units (NTU). According to an example embodiment, the turbidity property can be used to estimate the content of insoluble matter inside, such as solution cleanliness and particle concentration. According to an example embodiment, it can also be used to estimate the time evolution of the insoluble matter in the droplet. According to an example embodiment, the statistics obtained from the turbidity can be used to track the evolution of chemical or biological reactions inside the droplet. Any one of these characteristics or a combination thereof can be used as a variable for adapting a control signal.

[0077] According to an example embodiment, a set of predetermined characteristic values related to the characteristics of the droplet (such as the volume and viscosity of the droplet, etc.) are stored as reference values with a margin. During operation, when the characteristic value exceeds the margin, these characteristics can be used to adapt subsequent control signals to fine-tune EWOD parameters, such as the voltage or frequency applied to the electrodes.

[0078] According to an exemplary embodiment, as Figure 5 shown, some properties or characteristics can be measured over a time interval. The property can be a property of a droplet, such as viscosity, shape, and volume. The property can also be an interaction between droplets, such as droplet stability. Thus, for example, the merging of similar or different media in the form of droplets or split droplets can be analyzed with high temporal and spatial resolution.

[0079] According to an exemplary embodiment, features inside the droplet are also captured in the image, such as an object or objects. According to an exemplary embodiment, the object can be, but is not limited to, a biological entity, such as a cell, a molecule, a molecular fragment, etc. A biological cell refers to a structured and functional unit of a life form, such as a cell formed by cytoplasm enclosed in a membrane, which contains many biomolecules, such as proteins and nucleic acids. A molecule refers to a biomolecule, such as RNA, DNA, protein, etc. A molecular fragment can be an RNA fragment, a DNA fragment, a peptide, etc. According to an exemplary embodiment, properties of a biological cell can be captured in the image, such as size, shape, spectral absorptivity, spectral transmittance, etc. According to an exemplary embodiment, properties of a biological cell in the image can be captured over a time interval, such as cell growth and development, etc. According to an exemplary embodiment, when in response to a biological entity, properties of a biological cell can be captured in the image, the biological entity such as a cell that reacts to a certain chemical substance, an intercellular reaction, etc.

[0080] As Figure 7 shown, according to an exemplary embodiment, unexpected objects such as bubbles, micelles, or other liquid-phase irregularities are captured in the image. The unexpected object may indicate that the electrode in contact with the droplet is not working properly. The electrode may fail in the future. According to an exemplary embodiment, the control signal is adapted to send the droplet currently in contact with the electrode to an adjacent electrode. According to an exemplary embodiment, the control signal is adapted to avoid using this electrode to replace other droplets. Any designed path of other droplets will be recalculated and redesigned. In addition, the electrode can be turned off by the control unit (104).

[0081] According to an exemplary embodiment, the object can be another droplet. According to an exemplary embodiment, the control signal can be adapted to manipulate the droplet based on the difference in properties of two droplets captured in the image. According to an exemplary embodiment, the control signal can be adapted to manipulate the droplet based on the change in properties of two droplets captured in the image over a time interval.

[0082] According to an exemplary embodiment, the EWOD chip includes a plurality of EWOD layers. Each layer functions independently and can be controlled individually and in parallel by a control unit (104).

[0083] According to an exemplary embodiment, a TFT backplane, an EWOD layer, and a glass cover form an EWOD sub-operation unit, where a plurality of EWOD sub-operation units are stacked on top of each other. According to an exemplary embodiment, the control unit (104) may have a sub-control module for controlling each EWOD sub-operation unit. The lensless imaging device (31) can image through the plurality of stacked EWOD sub-operation units without mechanical focusing or movement. Focus adjustment is digitally completed from the recorded hologram to focus on multiple layers simultaneously. The control signal is individually adapted to the droplets in each EWOD layer.

[0084] According to an exemplary embodiment, the preferred arrangement of a double-layer EWOD chip is as Figure 8As shown. The EWOD chip includes a glass cover (24) and a first EWOD layer (251), with a first operating space formed therebetween. The first TFT backplane (261) is coupled to the first EWOD layer (251). The EWOD chip further includes a second EWOD layer (252), with a second operating space formed between the second EWOD layer (252) and the glass cover (24). The EWOD chip also includes first and second carriers (221, 222) for supporting the respective first and second TFT backplanes (261, 262), and first and second electronic interfaces (211, 212) for receiving control signals from the drive module of the instrument (100). The first and second electronic interfaces (211, 212) are electrically coupled to the respective first and second TFT backplanes. The jet (25) can be shared for supplying fluid to the first and second operating spaces. In operation, the light source (32) emits light onto the EWOD chip, forming an illumination area on the EWOD chip. The lensless imaging device in the droplet detection system (103) includes an imager chip (312) and an imager PCB (311), and the imager PCB detects light and captures a holographic image. The control unit (104) receives the image from the droplet detection system (103) and adapts the control signal to be sent to the drive module based on the received image. When an image of a droplet in the first operating space is taken, the adapted control signal based on the image is sent to the first interface (211). When an image of a droplet in the second operating space is taken, the adapted control signal based on the image is sent to the second interface (211). According to an example embodiment, the two EWOD layers will share a common ground electrode. According to an example embodiment, the respective TFT backplanes for the two EWOD layers are driven by two different controllers, i.e., driven in parallel and independently in the control unit (104). According to an alternative example embodiment, they can be driven by a single controller in a time-division multiplexing manner, i.e., a time-switching manner. This helps to reduce costs.

[0085] According to an example embodiment, the control unit (104) sends a control signal to the droplet detection system (103) to instruct the lensless imaging device (31) to scan in a region or sub-region within the illumination area.

[0086] According to an example embodiment, as Figure 9 shown, the droplet detection system (103) further includes fluorescence detectors (FLO1 and FLO2). Fluorescence detection requires a fluorescence light source. According to an example embodiment, the light source (32) includes a first sub-light source for lensless imaging and a second sub-light source for fluorescence detection. The light source (32) may further include a dichroic mirror such that the first sub-light source is transmitted through the first side of the mirror onto the EWOD chip, and the second sub-light source is reflected onto the EWOD chip at the second side of the mirror.

[0087] In a third aspect, the present invention relates to a droplet manipulation method. According to an example embodiment, at least two droplets with adapted control signals are manipulated simultaneously. The control signal for each of the at least two droplets is adapted according to at least one received image. According to an exemplary embodiment, at least one hundred droplets are manipulated by image-based adapted control signals. According to an exemplary embodiment, at least one thousand droplets are manipulated by image-based adapted control signals. According to an exemplary embodiment, at least one million droplets are manipulated by image-based adapted control signals.

[0088] According to an example embodiment, the control unit (104) assigns assays to each droplet. An image is taken at each step of the assay. According to an example embodiment, a plurality of droplets are transferred to a predetermined position. The path of each droplet is designed individually and globally. According to an example embodiment, the paths of the plurality of droplets are calculated such that: 1) the number of electrodes reused is minimized, and 2) the total time of displacement of the plurality of droplets is minimized. The efficiency of droplet transfer is maximized, and premature electrode degradation due to overuse in a short time, i.e., hysteresis and permanent charging, is avoided simultaneously.

[0089] According to an example embodiment, as a first step, the position of each droplet is captured in the image, and the target position of the droplet is predetermined. As a second step, the movement path of each droplet in the droplet group is designed. As a third step, the total movement time, the number of electrodes reused, and the number of times each electrode is reused are calculated. The second and third steps are iteratively repeated for different movement paths of each droplet, and thus the time and electrode reuse information can change during the iteration. When the desired path strategy is achieved, for example, when the time and electrode reuse situation no longer improves, the repetition ends.

[0090] According to an example embodiment, the control unit (104) is configured to generate droplets including objects. After a solution containing hundreds of thousands of objects in the range of about 1 - 100 microliters is loaded into the cartridge fluid, a predetermined area of the operating space in the EWOD chip is activated to further load the solution into several columns of the electrode array, as Figure 10 shown. Thus, there is a certain volume of solution on the EWOD chip. As Figure 11As shown, the unit cells of the m×n electrodes are activated to generate droplets from a predetermined region in which a solution containing an object is present. According to one exemplary embodiment, n is 3. The unit cells can be repeatedly activated on the EWOD layer in a column to perform parallel operations for droplet generation. One row of electrodes exists between the unit cells. In each such operation sequence, the generated droplets in each column are moved to the next column of electrodes, and new droplets are generated in a column. Thus, more than one droplet can be generated in parallel. According to an exemplary embodiment, the unit cells for droplet generation can be changed to different electrodes to prevent electrode wear due to repeated actions of droplet generation. For a practical design with a row size of 400 electrodes and a frame rate of 10 Hz, it is feasible to generate hundreds of individual droplets per second from one inlet of the cartridge. Hundreds of thousands of individual droplets can be generated on the EWOD chip within a few minutes.

[0091] Some defects on the EWOD chip are not visible in the captured images. Utilizing the electro-optic effect in the droplets, the system can be used for quality inspection of the EWOD chip, especially suitable for invisible defects. According to one exemplary embodiment, in operation, at least one droplet containing an electro-optic material sensitive to an electric field is introduced into the EWOD chip. When the EWOD electrode in contact with the droplet is activated, the contact angle between the EWOD chip and the droplet changes. Through a lensless imaging device, the change in the effective dielectric constant will result in a change in the holographic pattern of the droplet. The relative change in the dielectric constant can be used as a characteristic for testing whether the EWOD electrode is operating within the specification. If the relative change in the dielectric constant is not within a predetermined margin, the subsequent control signals for all droplets will be adapted to avoid using the said electrode. This is an advantage as it eliminates the need for an additional inductive sensing circuit for detecting hardware problems. The gain in circuit area can be used for other useful features, such as larger capacitors for faster droplet actuation. Another advantage is that this test can be used periodically to observe the evolution of circuit reliability using a single or a few images. A third advantage is that it can be read quickly and data can be easily obtained. No additional hardware complexity is required. According to one exemplary embodiment, the quality inspection can be performed before manipulating the droplets of interest. According to one exemplary embodiment, the quality inspection can be performed during the manipulation of the droplets of interest.

[0092] In a fourth and fifth aspect, the present invention also relates to a computer program and a computer-readable medium storing thereon. The computer program executes the described method in an instrument (100) or system. The instrument (100) saves all experimental data and analysis results of the experimental operation of the droplets to a local disk or uploads them to cloud storage. According to an example embodiment, the instrument (100) further includes a user interface that notifies the user of the progress of the experimental operation, instructs the user to follow certain steps, and receives inputs from the user related to experimental details. The user can obtain all available information for each experiment in a timely manner.

Claims

1. An instrument (100) for droplet manipulation, comprising: A driving module (101) configured to receive a control signal and operate an electrowetting-on-dielectric chip (102) with an operating medium, wherein the electrowetting-on-dielectric chip (102) is configured to manipulate droplets (1); A droplet detection system (103) comprising a light source (32) and a lensless imaging device (31), the lensless imaging device being configured to obtain at least one image including the droplet (1) in the electrowetting-on-dielectric chip (102); A control unit (104) configured to receive the at least one image from the droplet detection system (103) and to send a control signal to the driving module (101); Wherein the control unit (104) is further configured to adapt at least one of the control signals based on the at least one image.

2. A system comprising the instrument (100) according to any one of the preceding claims, further comprising an electrowetting-on-dielectric chip (102), preferably in a replaceable cartridge, configured to receive a signal from the driving module (101).

3. The system according to claim 2, wherein The at least one adapted control signal is for instructing the driving module (101) to move the droplet to a predetermined position.

4. The system according to any one of claims 2 to 3, characterized in that Wherein the control unit (104) is configured to extract the position of the droplet from the at least one image, and wherein the at least one control signal is adapted based on the position.

5. The system according to claim 4, characterized in that, The control unit (104) is configured to adapt the control signal such that if the movement of the droplet after a first control signal is different from a predetermined first path, at least one subsequent control signal is sent to move the droplet along a second path different from the first path to the predetermined position.

6. The system according to any one of claims 2 to 5, characterized in that, The at least one image includes a holographic image.

7. The system according to any one of claims 2 to 6, characterized in that The at least one image includes a two-dimensional or three-dimensional image.

8. The system according to any one of claims 2 to 7, characterized in that, The control unit (104) is configured to adapt at least one of the control signals based on at least one variable obtained by the control unit (104) from the at least one image, each of the at least one variable representing a different property selected from one or more of size, shape, spectral absorptivity, spectral transmittance, turbidity, viscosity, and the spatial distribution of at least one of the foregoing properties of the droplet.

9. The system according to any one of claims 2 to 8, characterized in that, The at least one control signal is adapted to instruct the driving module (101) to mix at least two droplets into one droplet; and optionally, the control unit (104) is configured to adapt at least one of the control signals based on at least one variable obtained by the control unit (104) from the at least one image to continue or stop the mixing of the droplets, the at least one variable representing the spatial distribution of at least one property or combination of properties selected from one or more of size, shape, spectral absorptivity, spectral transmittance, turbidity, and viscosity of the droplet.

10. The system according to any one of claims 2 to 9, characterized in that, The control unit (104) is configured to adapt at least one of the control signals based on at least one variable obtained by the control unit (104) from the at least one image, each of the at least one variables representing an object in the droplet.

11. The system according to claim 10, wherein The object is a cell, a bubble, or a different immiscible droplet.

12. The system according to any one of claims 2 to 11, characterized in that, The control unit (104) is configured to adapt at least one control signal based on at least one variable obtained by the control unit from the at least one image, each of the at least one variables representing an object in the droplet.

13. The system according to claim 6, wherein, The dielectric electro-wetting chip (102) includes a plurality of dielectric electro-wetting layers, wherein the control unit (104) is configured to adapt at least one of the control signals based on at least one variable obtained by the control unit (104) from a plurality of holographic images by using focus adjustment, each of the at least one variables representing the presence or absence of a droplet in a different dielectric electro-wetting layer.

14. A method for droplet manipulation in an instrument according to claim 1, comprising the steps of: a. causing the control unit (104) to receive at least one image including a plurality of droplets; b. causing the control unit (104) to generate control signals for simultaneously manipulating the droplets by electro-wetting based on the received at least one image.

15. A computer program comprising instructions for causing the instrument according to claim 1 to perform the steps of the method according to claim 14.

16. A computer-readable storage medium having stored thereon the computer program according to claim 15.

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