Techniques for displacing deposits from a particle sorter

Abstract

A particle sorter 1 comprises a fluid channel 10 and a bubble generator 12, wherein bubbles generated may displace deposits in the particle sorter. The deposits (e.g. dirt, debris, gunk) may be downstream or on the bubble generator (figure 5). The bubble generator may be a microheater, acoustic actuator or a focused laser, wherein the bubbles may be generator by mediating the voltage pulses. The build up of deposits may be detected by machine vision to determine when bubbles should be generated to clean the channel. The particle sorter may comprise a single-junction sorter comprising an inlet 11, an output sort channel 14 and an output waste channel 13. The bubble generator may be operated to displace fluid around a particle to be sorted to create a transient flow to direct the particle. There may be a vortex element present to cause a vortex in the transient flow to direct the particle.

Description

FIELD OF THE INVENTION The invention relates to techniques for displacing deposits from a particle sorter. BACKGROUND Instruments for particle sorting, also known as particle sorters or cell sorters, are widely used in biological research, clinical diagnostics, and therapeutic applications to isolate cells, viruses, organelles, encapsulated cells, exosomes, or synthetic beads from heterogeneous suspensions in a fluid medium. The most widespread particle sorting technology is based on the electrostatic deflection of droplets in air (conventional particle sorters). Recently, microfluidic cell sorters have been developed and are gaining prominence. Particle sorters generally align particles into a single file within a fluid suspension. They then detect and identify these particles and separate the target particles from the remainder of the particles in a sample. Conventional particle sorters use sheath flow focusing, also known as hydrodynamic focusing, which injects particles into a core stream surrounded by a concentric stream of a sheath fluid. The flow is then constricted to increase the particle velocity, giving a greater separation between particles and aligning them more tightly to the centre of the channel. The combined stream enters an optical cuvette or nozzle where detection occurs. Here, the particles are illuminated by one or more focused laser beams where the interaction scatters light and emits fluorescence (if the particles are labelled with fluorescent markers). Detectors and electronics collect this light and quantify the transient signal, reporting height, areas, and peaks widths of the peaks at each detector channel output. These measurements are displayed on a computer in cytometry plots, where one or more gates can be drawn to select a target population based on the magnitude of the scattered light and fluorescence. The gates are uploaded to the instrument to make real-time sorting decisions. Following detection, the particle flow leaves the sorter through a nozzle (with a diameter typically in the range of 50- 150 pm), where break-up into droplets is stimulated mechanically to create a uniform droplet stream. If the droplet contains a target cell, an electric charge is applied at the moment of droplet break-off. Subsequently, an electrostatic field deflects the droplet into a collection vessel or the waste stream. The amount of charge on the droplet determines the magnitude of deflection and into which vessel it is collected. All particle sorters, including conventional particle sorters and microfluidic particle sorters, rely on the flow of the fluid or suspension through narrow channels or orifices and are therefore prone to clogging, i.e. blockages caused by “gunk”, which includes cell aggregates, debris build-up, or precipitates. Other physical impediments to sorting that can build up in a narrow channel or orifice are trapped gas bubbles and the build-up of biomolecule fouling films such as protein films. These can compromise sorting performance, end sorting prematurely, or damage the instrument such that it requires servicing or repair. Moreover, particle sorters may also have issues with trapped gas bubbles in the flow channels. The source of gas bubbles can either be residual air from the filling and wetting of the dry channel or outgassing from a fluid. Trapped bubbles can adversely affect the cytometry measurements, for example, by interacting with the laser beam. Trapped bubbles may also jeopardise the sorting deflection, for example, by absorbing mechanical energy from the actuator in either a conventional particle sorter or a microfluidic particle sorter. Additionally, particle sorters may be sensitive to the build-up of a biomolecule film on the actuator or other contacting surfaces. Examples include microfluidic particle sorters with an embedded actuator, such as surface acoustic wave particle sorters, or particle sorters based on a thermal inkjet-like actuator, such as the vortex-actuated particle sorter. In the latter, the actuator is embedded in the channel and uses rapid heating to form a thermal vapour bubble. The localised heat from the actuator can cause biomolecules to denature and adhere to the surface, creating a film. As the film grows, it acts as an insulating layer, reducing heat transfer into the liquid. This requires the actuator to be driven at higher voltage to maintain the optimum deflection of particles. Eventually the film growth is sufficient that the actuator can no longer provide enough energy to the bubble, compromising sorting performance. Therefore, a significant issue for all particle sorters is the clogging of channels or nozzles. At best, this can impair sorting performance; at worst, it can prevent sorting and even require resolution by a service engineer. These blockages, often caused by deposits (or “gunk”), usually occur where the flow is constricted. For droplet sorters, this is often in the nozzle or optical cuvette. In microfluidic sorters, this is typically at narrow points in the channel or stagnation points, such as at channel junctions. A current option to reduce the probability or frequency of clogging is to reduce the formation of aggregates and debris by altering the preparation of samples. Several sample preparation methods can reduce clogging. Particles may be passed through a strainer before being loaded to remove aggregates of particles larger than the channel width or nozzle diameter. The sample in the input vessel may be agitated (e.g. vortex mixing or magnetic stirring) to prevent particles from aggregating. Certain additives in the fluid suspension may reduce clogging by reducing aggregation of components that constitute deposits or “gunk” such as cells, debris and biomolecules. These include serum, serum proteins, such as bovine serum albumin (BSA), surfactants such as Pluronic F-68, or enzymes such as DNase. Regular maintenance and cleaning of the fluidic system is recommended for conventional sorters, and these instruments often have selfcleaning protocols. However, none of these methods completely prevent clogging, and some may be incompatible with the sample or the downstream application of the particles. Further, if clogging occurs during the sorting process, the sorting process must be interrupted and the blockage cleared. Stopping the sorting process of any particle sorter is undesirable and may mean the sorted sample is compromised or lost. Cleaning solutions with detergents or enzymes may be used to dissolve or dislodge the gunk. If the blockage is still not removed, the channel, nozzle or cuvette may need to be replaced or manually washed out. Some particle sorters may require attendance by a service engineer to replace the required parts. For example, microfluidic particle sorters process particles in microfluidic channels, typically as part of a cartridge with connecting tubes for one sample input and two or more sample outputs. While the detection of the particles may be similar to conventional particle sorters, microfluidic particle sorters may align and deflect particles by different means. For alignment, some microfluidic particle sorters use hydrodynamic focusing to constrain the particle in one or two dimensions within the fluidic channel cross-section; others use inertial focusing, while others use narrow channels without any further particle alignment. Deflection mechanisms are also varied, but all involve a single microfluidic channel splitting into two or more channels following the detection point. If not actively deflected, particles will passively flow down a negative output or waste channels. When the actuator is activated, particles are deflected into streamlines flowing into one or more target output channels. Microfluidic particle sorters typically have disposable or single-use fluidic cartridges which can be replaced in order to provide a new cartridge free from deposits. In this way, microfluidic devices require regular maintenance and cleaning to maintain sorting performance. The invention disclosed herein provides a solution to clear clogging, air bubbles and fouling films from the narrow channels or orifices of a particle sorter without interrupting the sample flow or disconnecting the fluidic system. SUMMARY OF INVENTION According to a first aspect of the invention a method for displacing deposits from a particle sorter comprising a fluid channel and bubble generator is provided, the method comprising: providing the fluid channel with a flow of a fluid; and selectively operating the bubble generator in order to transfer energy to the fluid so as to generate a plurality of bubbles in the fluid such that the plurality of bubbles displace one or more deposits downstream of the bubble generator. Particle sorters receive a flow of a particle-containing fluid through a fluid channel and are therefore prone to clogging i.e. blockages. As used herein, the word “deposits” (or “gunk”) encompasses various impediments that may lead to this clogging, including but not limited to one or more of: debris build-up or aggregates in the fluid channel, trapped bubbles in the fluid channel, and / or biomolecule films on an inner walls of the fluid channel. The debris may include aggregated cells or cell clumps, dust, impurities or precipitates. The biomolecule films may form at various regions of the inner walls of the fluid channel, but are particularly susceptible to formation in the region of components configured to actuate the sorting of particles. Difficulties arise when removing these deposits as present solutions require the sample flow to be interrupted and components of the device to be disconnected so as to manually wash out or replace the fluid channel. Therefore, by operating a bubble generator to selectively transfer energy to the particle-containing fluid, nucleation of thermal vapour bubbles occurs in the particle-containing fluid as gas pockets are formed in the fluid. The plurality of bubbles travel downstream in the fluid channel. The heating and cooling, rapid changes in pressure, fluctuations in shear forces, and surface tension of the vapour-fluid interfaces all cause mechanical agitation which can break down and dislodge the deposits as the bubbles flow past. Advantageously, the deposits can be removed from the fluid channel without requiring interruption to the flow of fluid. The bubble generator may be operated using a control system. Particle sorters are used to isolate specific cells or particles. The particle sorter may be a conventional particle sorter or a microfluidic particle sorter. The bubble generator may be placed in a sample, sheath, or a buffer inlet upstream of the fluid channel. In some examples, the fluid channel comprises at least a microfluidic channel, a nozzle or a junction. As used herein, the word “particle” encompasses biological cells, solid beads, and liquid droplets of one liquid phase in a carrier fluid (such as aqueous droplets in a non-aqueous carrier fluid). Liquid droplets may themselves contain further particles. As used herein, the word “fluid” encompasses both aqueous and non-aqueous fluids, typically in the liquid or gas phase. In some examples, the fluid is a particle containing fluid, wherein the particle-containing fluid preferably comprises particles to be sorted. In this way, the method can comprise displacing deposits from the particle sorter during a sorting process (i.e. during use), rather than halting the flow of the fluid containing the particles to be sorted and then using a further fluid to displace the deposits. However, fluids not containing particles may also be used, for example, to displace deposits between uses of the particle sorter. The fluid may be any liquid that is sufficiently volatile for the bubble generator to generate a bubble, such as water, an aqueous solution, or a non-aqueous carrier medium. The skilled person will understand that the terms “particle” and “fluid” are not limited to the above definitions should also be interpreted according to their understood meanings in the art. The fluid channel of the particle sorter preferably comprises one or more of a narrow (microfluidic) channel or an orifice. The size and shape of the fluid channel of the particle sorter may be selected based on the intended application, that is that characteristics (e.g. length and / or width) of the fluid channel may be based on the particle type desired to be isolated. As used herein, the term “narrow” refers to a fluid channel having a channel width in the range 0 - 1 millimeters, and preferably in the range 20 - 200 micrometers. I n some examples, the fluid channel has a channel cross section of 40 x 70 micrometers, although other cross section dimensions may be contemplated. In some examples, operating the bubble generator comprises applying a pulse chain to the bubble generator, the pulse chain comprising a sequence of M series of N pulses. In some examples, N is an integer value greater than one. In some examples, each pulse causes the homogeneous nucleation of a thermal vapour bubble. As such, the series of pulses generates a bubble stream in the fluid which provides further agitation to the deposit(s). In some examples, M is an integer value greater than or equal to one. In this way, a bubble stream comprising an increased number of bubbles may be generated when M is increased, thereby providing further agitation to break down and / or dislodge the deposit. In some examples, applying each series of N pulses comprises alternating between operating the bubble generator at a first power dissipation setting for a first time duration and operating the bubble generator at a second power dissipation setting at a second time duration. In some examples, the ratio of the first time duration to the second time duration may vary when applying the series of N voltage pulses. Operating the bubble generator at a first power dissipation setting results in a dissipation of a first amount of energy to the fluid. Operating the bubble generator at a second power dissipation setting results in a dissipation of a second amount of energy to the fluid. In this way, the amount of energy transferred to the fluid is controlled by the power dissipation settings. There are various ways in which the transfer energy, or heat, to the fluid can be controlled. In some examples, operating the bubble generator at a first power dissipation setting may comprise applying a signal to the bubble generator having a pulse-width modulation (PWM) signal with a given ratio of on signal to off signal which results in controlled dissipation of energy to the fluid. In some examples, applying the pulse chain comprises intermittently applying the M series of pulses. In other words, the pulse chain comprises a gap between each pulse series. In this way, the bubble operator generates a bubble stream comprising an interruption in the bubble stream. The pulse chain provides further agitation to the deposit by the repeated rapid transitions between bubbles and liquid in the downstream flow. Further, by interrupting the bubble stream, the microheater may cool as the fluid passes over it. In some examples, the series of N pulses are separated by a third time duration. In this way, the bubble operator generates a bubble stream comprises a periodic interruption in the bubble stream. The periodic interruption has the duration of the third time duration. In some examples, controlling the heat transfer to the fluid comprises operating a voltage applied to the bubble operator. In some examples, the pulse chain comprises a voltage pulse chain, and wherein operating the bubble generator at a first power dissipation setting comprises applying a first voltage to the bubble generator and operating the bubble generator at a second power dissipation setting comprises applying a second voltage to the bubble generator. In some examples, one of the first voltage or second voltage is zero volts. In some examples, operating the bubble generator comprises adjusting one or more of: a value of M, a value of N, the first time duration, the second time duration, and the third time duration. Characteristics of the pulse chain determine the effectiveness of bubble stream at displacing and agitating varying types of deposits, e.g. air bubbles, debris, aggregates, and protein films. Therefore, by adjusting the above-mentioned characteristics of the bubble generator, the resulting characteristics of the bubble stream generated may be based on the deposit it is intended to agitate and displace. In this way, the displacement of deposits from a particle sorter is further improved. In examples where M=1 a continuous series of N voltage pulses is provided to the bubble generator, thereby generating a continuous bubble stream. In examples where M>1, series of A / voltage pulses are applied at intervals, such that N streams of bubbles are generated, each separated by the third time duration. In other words, there is a periodic interruption in the bubble stream. In this way, M can be selected based on the deposit intended to be agitated and displaced. For example, the value of M may be greater for displacing aggregates than the value of Mfor displacing trapped bubbles. In this way, the displacement of deposits from a particle sorter is further improved. In some examples, the method further comprises detecting one or more of said deposits in the particle sorter and operating the bubble generator based on the detected one or more deposits. In this way, the bubble generator may be automatically operated based on this detection and / or directly after this detection, so as to allow for automatic displacement of deposits in response to said detection. In this way, the device may only be operated to displace a deposit once one is detected, thereby conserving energy. Further, the bubble stream generated may be based on the deposit it is intended to agitate and displace. In some examples, detecting one or more of said deposits further comprises identifying one or more deposit types. As used herein, the term “deposit type” refers to classification of deposit based on its characteristics. Examples of deposit types include debris build-up or aggregates in the fluid channel, trapped bubbles in the fluid channel, and / or biomolecule films an inner wall of the fluid channel. Advantageously, by identifying a deposit type, the operation of the bubble generator can be optimised based on the identified deposit type. For example, a bubble trapped in the fluid may be removed using a different operating functions to that of the build-up of debris. This method provides precise and efficient displacement of deposits. In some examples, detecting one or more of said deposits further comprises monitoring an actuation voltage of the bubble generator to identify at least one deposit on the bubble generator. The actuation voltage is the voltage required to generate one or more bubbles are used to displace deposits and / or, in some embodiments, to initiate sorting of particles. Therefore, an actuation voltage is needed to maintain correct deflection of the target particle in the sorting process. As the heater fouls and develops a fouling film, it becomes increasingly thermally insulated from the fluid, thus requiring a larger energy transfer and hence a larger actuation voltage to maintain the particle deflection. Therefore, by monitoring the actuation voltage, it can be deduced when a fouling film or other deposit has developed on the bubble generator. In some examples, the method further comprises comparing the actuation voltage to a threshold voltage and identifying that the at least one deposit is on the bubble operator when the actuation voltage is above the threshold voltage. Once the actuation voltage increases to above a certain threshold, it can be deduced when a fouling film or other deposits has developed on the bubble generator, such that the bubble generator should be operated so as to displace one or more deposits downstream of the bubble generator. In some examples, detecting one or more of said deposits comprises imaging the fluid channel. Preferably, the imaging field covers at least a portion of the fluid channel and / or sorting region of the fluid channel (when applicable). Image processing algorithms can be applied to improve the detection of one or more said deposits. The detection of one or more of said deposits may comprise at least a location in the fluid channel, a flow pattern of particles or deposits in the channel, a density of one or more said deposits, a distribution of one or more said deposits etc. In some examples, imaging the fluid channel comprises using machine vision to identify at least one deposit in the particle sorter. Machine vision can rapidly analyse the images of the particle sorter to precisely identify at least one deposit in the particle sorter. In some examples, the particle sorter further comprises a cooling element, and wherein the method further comprises operating the cooling element. The cooling element may be configured to remove heat from the fluid or particle sorter. In this way, an excess of bubbles is prevented from being generating when the fluid gets too hot thereby maintaining a controlled bubble stream. Further, in some examples, the particles in the fluid may become damaged or denatured at too high of a temperature. Cooling the fluid or particle sorter advantageously prevents this. In some examples, the particle sorter comprises a single-junction sorter, wherein the fluid channel comprises an inlet channel, one or more output sort channels and an output waste channel, and wherein each of the one or more output sort channels and the output waste channel are connected to the input channel for receiving the fluid therefrom. In some examples, the method further comprising selectively operating the bubble generator to displace the fluid around a particle to be sorted and thereby creating a transient flow of the fluid in the input channel so as to direct the particle to be sorted into one of the output sort channels. In some examples, the particle sorter further comprises a vortex element configured to cause a vortex in the transient flow in order to direct the particle to be sorted into one or more output sort channels. In some examples, operating the bubble generator comprises operating the bubble generator during sorting of particles. According to a second aspect of the invention there is provided a particle sorter comprising a fluid channel configured to conduct a flow of fluid; the particle sorter further comprising a bubble generator operable to selectively transfer energy to the fluid so as to generate a plurality of bubbles in the fluid; and wherein the bubble generator is arranged such that the plurality of bubbles displace one or more deposits downstream of the bubble generator. In some examples, the particle sorter is a single-junction sorter, wherein the fluid channel comprises an inlet channel, one or more output sort channels and an output waste channel, wherein each of the output sort channel and the output waste channel are connected to the input channel for receiving the fluid therefrom; and the bubble generator is further operable to selectively displace the fluid around a particle to be sorted and thereby to create a transient flow of the fluid in the input channel; and the particle sorter further comprises a vortex element configured to cause a vortex in the transient flow in order to direct the particle to be sorted into one or more output sort channels. In some examples of the first and second aspects of the invention, the bubble generator comprises one or more of: a microheater, an acoustic actuator, and a laser focused towards the fluid channel. The microheater may be operated such that heat is applied to the fluid to induce nucleation of vapour bubbles in the fluid. An acoustic actuator may be operated such that acoustic waves are applied to and absorbed by the fluid to induce nucleation of vapour bubbles in the fluid. A laser may be focused on the fluid, wherein the fluid is an absorbing medium. In some examples, the microheater comprises a stacked arrangement comprising one of more of: a base layer, thermal buffer layer, an electrically insulating passivation layer, an anti-cavitation layer, an interface layer. In some examples, the particle sorter further comprises an adjustable voltage regulator coupled to the bubble generator, the adjustable voltage regulator configured to apply a voltage pulse chain to the bubble generator, the voltage pulse chain comprising a sequence of M series of N voltage pulses. In some examples, the particle sorter further comprises a cooling element. The particle sorter provided according to the second aspect of the invention may also be configured in any of the manners described above in relation to the first aspect of the invention. According to a third aspect of the invention there is provided a bubble generator for use with a particle sorter comprising a fluid channel configured to conduct a flow of fluid, the bubble generator being operable to selectively transfer energy to fluid so as to generate a plurality of bubbles in the fluid, wherein the bubble generator is arranged such that the plurality of bubbles displace one or more deposits downstream of the bubble generator. The bubble generator may be configured in any of the manners described above in relation to the first and second aspects of the invention. BRIEF DESCRIPTION OF DRAWINGS Figure 1 shows a schematic of a particle sorter according to an embodiment of the present invention; Figure 2 shows a schematic of a microheater according to an embodiment of the present invention; Figure 3 shows a schematic of an exemplary voltage pulse chain used to drive the bubble generator in an embodiment of the present invention; Figure 4 shows an exemplary electronic drive architecture used to drive the bubble generator in an embodiment of the present invention; Figures 5a to 5c show schematics of a particle sorter according to embodiments of the present invention wherein exemplary deposits are depicted in and / or on the particle sorter; Figure 6 shows a schematic of a particle sorter according to an embodiment of the present invention, wherein the particle sorter is further configured so as to direct the particle to be sorted into an output sort channel; Figure 7 shows a schematic of the variation over time of an actuation voltage of the bubble generator during an auto defouling method in an embodiment of the present invention; Figure 8 shows experimental data of the variation overtime of an actuation voltage of the bubble generator during an auto defouling method in an embodiment of the present invention; Figures 9a to 9c show experimental data of an auto defouling method according to an embodiment of the present invention; Figures 10a to 10c show experimental data of a power flow clear method on the junction region according to an embodiment of the present invention; Figure 11a to 11c show experimental data of power flow clear method on a constriction in the flow channel according to an embodiment of the present invention. DETAILED DESCRIPTION A particle sorter 1 is shown schematically in Figure 1. The particle sorter 1 comprises a fluid channel 10 comprising an inlet channel 11 leading to a first sort channel 13 for non-target particles and a second sort channel 14 for sorting target particles. The bubble generator 20, which in this example is a microheater 20, sits inside a heater pocket adjacent to the inlet channel 11. The microheater 20 is connected to an instrument through electrical tracks 24. The sorting tip 12 follows the heater pocket, where the channel narrows slightly, and allows for vortex-actuated particle sorting. In operation, a pulse of electricity is applied to the microheater 20, sufficient to cause the nucleation and expansion of a thermal vapour bubble, and is timed precisely with the passage of a target particle. The transient fluid displacement caused by the thermal vapour bubble in turn causes the generation of a vortex at the sorting tip 12. The vortex flows downstream with the target particle, deflecting the particle across the streamlines into the sort channel 14. This embodiment comprises a particle sorterwith a single microheater 20 (also referred to as a microresistor) for both actuation of a sort event and generation of bubbles for displacing deposits. However, alternative embodiments which include alternative types of particle sorters may comprise a separate bubble generator to generate bubbles for the same purposes. In some examples, the microresistor 20 may be made with microelectrical and micromechanical systems (MEMS) processing techniques. The microheater may comprise a stacked arrangement (see Figure 2). In this example, the microheater 20 comprises a silicon wafer as the base 26 with a thermal oxide acting as short time scale thermal buffer 25. Tantalum Nitride may be used as an electrically resistive layer 24 which may be protected with a Silicon Nitride electrically insulating passivation layer 23. The heater area may then be covered with a Tantalum anti-cavitation passivation layer 22 to protect from damage from cavitation bubbles. The microresistor 20 may have aluminium copper electrical tracks 24 that travel outside the channel terminating at electrical contacts used to interface with the main instrument electronics. Resistive heating occurs when a potential difference is applied to the electrical contacts. As measured by the instrument, the electrical resistance may be determined by the sum of the electrical track resistance and the micro resistor 20. Ina specific example, this is 250 total resistance with 12.50 across the tracks and 12.50 across the microresistor 20. In some implementations, such as shown in Figure 1, the surface area of the microresistor 20 may affect not only the fluid contact area and volume of fluid heated but also the power dissipation per area and thus the heating rate. Consequently, this surface area may be varied to improve the operational characteristics of the microresistor. In a specific example, the area is 80pm x180pm. The fluid contact area may be made smaller or larger to suit the channel size. Described herein are several gunk-clearing functions (or “modes”) which the present invention can employ to clear different types of deposits or “gunk”. The principle of each gunk-clearing function is to drive the microheater 20 with a programmed number of voltage pulses at a specific voltage, as shown in Figure 3. The voltage pulse-on time (t_on), voltage pulse-off time (t_off), pulse-on voltage (V), and the number of pulses (N) can be optimised for each gunk clearing function. An additional function is to repeat the voltage pulse chain number of times (M) with a time gap (t_gap) between them. The general terms “deposits” and “gunk” are used interchangeably, herein, and can include the contribution of cells, gas, debris and biomolecule aggregates to clogging and fouling, specifically including the build-up of debris or aggregates that could block the channel, trapped bubbles in the flow channel that could interrupt particle detection or sorting, and biomolecule films on contacting surfaces that may impede an actuator. The terms “cells” and “particles” as described herein, and the terms “cell sorting” and “particle sorting” as described herein are used interchangeably. The individual voltage pulses to the heater 20 cause the homogeneous nucleation of a thermal vapour bubble. Typically, the bubble expands and collapses in a few microseconds; however, if repeated pulses are delivered in quick succession, enough heat may be transferred to the vapour that the bubbles can persist, grow and be shed by the heater, flowing downstream with the fluid medium. The heating and cooling, rapid changes in pressure, and surface tension of the vapour-fluid interfaces all cause mechanical agitation which can break down and dislodge gunk as the bubbles flow past. A series of pulses can be made into a pulse chain by repeating it one or more times with a gap between each pulse series. The pulse chain provides further agitation by the repeated rapid transitions from bubbles to liquid in the downstream flow. To deliver voltage pulses with precise timing, one option is to use an adjustable voltage regulator connected to the heater 20 circuit through a transistor. Figure 4 shows a configuration to drive the heater 20 and measure the circuit resistance. Here, an MCU configures the voltage regulator and the FPGA PWM settings and triggers the FPGA to start the pulse chain. Separately, there is a parallel circuit where the MCU can use the regulator voltage to drive a resistor in series in the heater. The voltage drop across the resistor can then be used to infer the electrical resistance of the heater circuit. A control system on an embedded computing device or separate computing device with a user interface decides whether to trigger gunk clearing and which function to run; it then starts gunk clearing by sending a message to the MCU. Described herein are three gunk-clearing functions relating to the particle sorter shown in Figure 1. A first gunk-clearing function, referred to as the “power flow clear function”, is designed to clear trapped debris. Figures 5a and 5b show example particle sorters 1 in which debris has become trapped in such a way that the power flow clear function is suitable to clear said debris. Specifically, Figure 5a shows debris trapped at the channel junctions, and Figure 5b shows aggregates clogged at constrictions or bends in the flow. The debris or aggregates can require robust mechanical agitation to remove them, so the heater 20 is driven with a pulse chain comprising several short bursts with gaps to give repeated bursts of shedding defouling bubbles. In a specific example, the settings are N=1000 pulses of V=17.5V, ton= 4ps, t0 / / =16ps, repeated M=40 times with tgap=20 ms. A second gunk-clearing function, referred to as the “debubble function”, is designed to clear trapped air or gas bubbles, for example those which have outgassed from a saturated medium on surface features or channel walls, often in regions with slower fluid flow. An example of a trapped air bubble is shown in Figure 5c. These tend to be easier to remove than the built-up debris or aggregates and the heater 20 can be driven with a lower voltage and fewer pulses. Preferably, the heater 20 is driven such that it the temperature of the local area is maintained below a temperature that would create more air bubbles. In a specific example, the settings are N=2500 pulses of V=16.0V, tow=4ps, t0 / / =16ps, with no repeats, i.e. M=1. A third gunk-clearing function, referred to as the “actuator defoul function”, is designed to clear biomolecule foul films from the heater 20. An exemplary film is shown in Figure 5d. In a specific example, the settings are N=50000 pulses of V=16.5V, tow=4ps, t0 / / =16ps, with no repeats, i.e. M=1.As would be understood by the skilled person, the particle sorter of Figure 1 includes trapped bubbles which are easier to clear than aggregates, hence the difference in operation described above. However, in other particle sorter designs the films may be easier to clear. Therefore, the above-mentioned three functions may be altered according to the design of the particle sorter. A further step of any of the methods described herein is to block flow into the target channel 10 whilst gunk clearing is taking place. This prevents non-target particles, debris, or aggregates from entering the target channel 12 and contaminating the sorted population. This channel should be closed before starting the gunk clearing voltage pulses and opened again once the voltage pulses end. In a microfluidic sorter 1, this can be done, for example, with a solenoid valve to clamp soft tubing taking flow from the target particle microfluidics channel to the collection vessels. The particle sorter and method may also measure the resistance of the heater circuit and adjust the clearing function voltage accordingly. This ensures the defouling bubbles are generated similarly across separate devices with varying resistances due to manufacturing tolerances. If the resistance is higher than the reference resistance and voltage is not corrected, then vapour bubbles may not be powerful enough to clear the gunk. If the resistance is lower than the reference resistance and the voltage is not corrected, the heater can be overdriven, potentially causing damage. As it is constant power that is required, the gunk clear / x 0.5 / p \ function voltage should be rectified as V = —J Vref, where Rref is the reference voltage, 25Q for an example heater, and Vref is the reference voltage for the clearing function, which may be, for example, 17.5V, 16V, and 16.5V for the power flow clear function, debubble function and actuator defoul function, respectively. The particle sorter 1 may also include an active cooling system that removes excess heat produced by the heater 20. Excess heat can cause outgassing of air from the fluid, reduce the heater lifetime, or give undesirable warming to the fluid or particle solution. To counter the heat input from the gunk clearing, the cooling system power should be boosted proportionally to additional heat energy from the chip from the gunk-clearing pulses. As the heater 20 is driven over a relatively short time scale, the cooling power is increased for a duration T, for example, such that Px^xMxNxtonx V2 / R, where M is the number of repeats of the pulse chain, N is the number of pulses in each pulse chain, ton is the on-time of each pulse, V2 is the on voltage of each pulse, and R is the resistance of the electrical tracks and heater 20. Preferably, T = 15 s. In some examples, before triggering gunk clearing, the gunk is detected. For different types of gunk, the detection method may vary. In some examples, machine vision is used to monitor debris or cell aggregates that build up or cause clogs at constrictions or channel junctions. The imaging field covers the sorting region and channel junction as shown schematically in Figure 6. The build-up of gunk is monitored at several points within this field of view. Once a threshold area is reached and the gunk has persisted over, for example, three or more image frames, power flow clear is triggered. Heater fouling can be tracked by monitoring the sort actuation voltage needed to maintain correct deflection of the target particle in the sorting process. Alternatively heater fouling can be tracked by monitoring an actuation voltage needed to generate bubbles in a gunk clearing function or mode. In some examples, the particle sorter 1 includes an automatic control system to maintain sorting, which functions as follows. Images of the sorter are acquired, as shown in Figure 6, using a strobe illumination for each frame. Each strobe illumination is at a given time lag from the particle detection that corresponds to the particle passing the junction and entering the sort 1 output channel 12. A machine vision algorithm analyses the resulting image for the particle's position: the deflection is the distance of the particle from the lower particle wall 15, and declension is the distance along the channel from the junction to the particle 16. The time lag is adjusted continuously so that the declension is fixed to a set distance. To maintain sorting, the deflection must be controlled to a set point, usually set as the centreline of the channel. A control loop is set up to adjust the sort actuation voltage until the deflection is at the set point. As the heater fouls, it becomes increasingly thermally insulated from the sample solution, thus requiring a larger energy transfer and hence a larger sort actuation voltage to maintain the particle deflection. Once the voltage increases by a certain threshold, actuator defoul is triggered to clear the heater 20. Figure 7 shows a schematic of the sort actuation voltage required to maintain sorting over time and the triggering of defouling. When sorting is turned on 31, the starting voltage is noted as the “fresh” sort actuation voltage. As sorting continues, the heater begins to foul, and the voltage required to main sorting increases 32. Once the difference between the current sorting voltage and the fresh starting voltage exceeds a threshold 33 the voltage is set to defoul voltage 34 and defoul is PWM is triggered. The defoul clears the gunk from the heater surface, and then sorting starts again from the fresh sort actuation voltage. Figures 8 - 11c show experimental results. In all these examples, liquid flow rate through the narrow channel was around 265 microlitres per minute. Figure 8 shows experimental results of auto-defoul. Each time the actuator voltage exceeds a threshold value of 19.7 V, the defoul function is triggered. After each actuator defoul, a lower voltage of around 18.5 V is required for sorting. In this experiment approximately 40 s elapses between defouls. Figures 9a to 9c shows experimental results of the actuator defoul function. Figures 9a, 9b, 9c shows the microheater 20 in operation. Before the defoul, the actuation bubble is small and does not cover the whole heater surface, as shown in Figure 9a, and as was shown schematically in Figure 5d. During the defoul, a stream of large bubbles is released from the microheater 20, as shown in Figure 9b. After the defoul, the actuation bubble again covered the majority of the microheater, as shown in Figure 9c. Figures 10a to 10c show experimental results of the power flow clear function on the junction region. Figures 10a and 10c show the junction in operation before and after the power flow clear, while Figure 10b shows the stream of bubbles being released by the power flow clear. Before the power flow clear, the junction has a large amount of debris trapped, as shown in Figure 10a, and as was shown schematically in Figure 5b. After the power flow clear, the junction is free of debris, as shown in Figure 10c. Figures 11a to 11c show experimental results of a power flow clear on a constriction in the flow channel. Figures 11a and 11c show the constriction in operation before and after the power flow clear, while Figure 11b shows the stream of bubbles being released by the power flow clear. Before the power flow clear, the constriction has been clogged by debris, as shown in Figure 11a, and as was shown schematically in Figure 5a. After the power flow clear, the junction is free of debris, as shown in Figure 11c. It will be understood that the invention has been described in relation to its preferred embodiments and may be modified in many different ways without departing from the scope of the invention as defined by the accompanying claims.

Claims

1. A method for displacing deposits from a particle sorter comprising a fluid channel and bubble generator, the method comprising:providing the fluid channel with a flow of a fluid; andselectively operating the bubble generator in order to transfer energy to the fluid so as to generate a plurality of bubbles in the fluid such that the plurality of bubbles displace one or more deposits downstream of the bubble generator.

2. A method according to claim 1, wherein the fluid is a particle-containing fluid, wherein the particle-containing fluid preferably comprises particles to be sorted.

3. A method according to any preceding claim, wherein operating the bubble generator comprises applying a pulse chain to the bubble generator, the pulse chain comprising a sequence of M series of N pulses.

4. A method according to claim 3, wherein N is an integer value greater than one.

5. A method according to claim 3 or claim 4, wherein M is an integer value greater than or equal to one.

6. A method according to any of claims 3 to 5, wherein applying each series of N pulses comprises alternating between operating the bubble generator at a first power dissipation setting for a first time duration and operating the bubble generator at a second power dissipation setting at a second time duration.

7. The method according to any of claims 3 to 6, wherein applying the pulse chain comprises intermittently applying the M series of pulses.

8. A method according to claim 7, wherein the series of N pulses are separated by a third time duration.

9. A method according to claims 3 to 8, wherein operating the bubble generator comprises adjusting one or more of: a value of M, a value of N, the first time duration, the second time duration, and the third time duration.

10. A method according to any of claims 3 to 9, wherein the pulse chain comprises a voltage pulse chain, wherein operating the bubble generator at a first power dissipation setting comprises applying a first voltage to the bubble generator and operating the bubble generator at a second power dissipation setting comprises applying a second voltage to the bubble generator.

11. A method according to claim 10, wherein one of the first voltage or second voltage is zero volts.

12. A method according to any preceding claim, wherein the method further comprises detecting one or more of said deposits in the particle sorter and operating the bubble generator based on the detected one or more deposits.

13. The method according to claim 12, wherein detecting one or more of said deposits further comprises identifying one or more deposit types.

14. A method according to claim 12 or claim 13, wherein detecting one or more of said deposits further comprises monitoring an actuation voltage of the bubble generator to identify at least one deposit on the bubble generator.

15. A method according to claim 14, the method further comprising comparingthe actuation voltage to a threshold voltage and identifying that the at least one deposit is on the bubble operator when the actuation voltage is above the threshold voltage.

16. A method according to any of claims 12 to 15, wherein detecting one or more of said deposits comprises imaging the fluid channel.

17. A method according to claim 16, wherein imaging the fluid channel comprises using machine vision to identify at least one deposit in the particle sorter.

18. A method according to any preceding claim, wherein the particle sorter further comprises a cooling element, and wherein the method further comprises operating the cooling element.

19. A method according to any preceding claim, wherein the particle sorter comprises a single-junction sorter, wherein the fluid channel comprises an inlet channel, one or more output sort channels and an output waste channel, and wherein each of the one or more output sort channels and the output waste channel are connected to the input channel for receiving the fluid therefrom.

20. A method according to claim 19, the method further comprising selectively operating the bubble generator to displace the fluid around a particle to be sorted and thereby creating a transient flow of the fluid in the input channel so as to direct the particle to be sorted into one or more output sort channels.

21. A method according to claim 20, wherein the particle sorter further comprises a vortex element configured to cause a vortex in the transient flow in order to direct the particle to be sorted into one or more output sort channels.

22. A particle sorter comprising a fluid channel configured to conduct a flow of fluid, the particle sorter further comprising:a bubble generator operable to selectively transfer energy to the fluid so as to generate a plurality of bubbles in the fluid; andwherein the bubble generator is arranged such that the plurality of bubbles displace one or more deposits downstream of the bubble generator.

23. A particle sorter according to claim 22, wherein the particle sorter is a single-junction sorter, wherein the fluid channel comprises an inlet channel, one or more output sort channels and an output waste channel, wherein each of the one or more output sort channels and the output waste channel are connected to the input channel for receiving the fluid therefrom; andthe bubble generator is further operable to selectively displace the fluid around a particle to be sorted and thereby to create a transient flow of the fluid in the input channel; andthe particle sorter further comprises a vortex element configured to cause a vortex in the transient flow in order to direct the particle to be sorted into one or more output sort channels.

24. A method according to any of claims 1 to 21 or a particle sorter according to claim 22 or claim 23, wherein the bubble generator comprises one or more of: a microheater, an acoustic actuator, and a laser focused towards the fluid channel.

25. A method according to any of claims 1 to 21 orclaim 24 ora particle sorter according to any of claims 22 to 24, wherein the microheater comprises a stacked arrangement comprising one of more of: a base layer, thermal buffer layer, an electrically insulating passivation layer, an anti-cavitation layer, an interface layer.

26. A method according to any of claims 1 to 21, claim 24, or claim 25 or a particle sorter according to any of claims 22 to 25, the particle sorter further comprising an adjustable voltage regulator coupled to the bubble generator, the adjustable voltage regulator configured to apply a voltage pulse chain to the bubble generator, the voltage pulse chain comprising a sequence of M series of N voltage pulses.

27. A bubble generator for use with a particle sorter comprising a fluid channelconfigured to conduct a flow of fluid, the bubble generator being operable to selectively transfer energy to fluid so as to generate a plurality of bubbles in the fluid, wherein the bubble generator is arranged such that the plurality of bubbles displace one or more deposits downstream of the bubble generator.

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