Noise reduction for an electrochemical sensor

Abstract

An electrochemical sensor apparatus having a sensor comprising a working electrode, a counter electrode, and optionally a reference electrode. The apparatus has a power source configured to apply a voltage to the sensor; and a voltage processor comprising one or more components. The voltage processor is configured to reduce a voltage perturbation in the voltage applied to the sensor.

Description

[0001] NOISE REDUCTION FOR AN ELECTROCHEMICAL SENSOR

[0002] FIELD

[0003]

[0001] . The present invention relates generally to electrochemical sensors, and particularly electrochemical aptamer-based sensors. The invention provides an improvement in the delivery of electrical power to electrochemical sensors allowing for more determination of target analyte concentration at low limits of detection.

[0004] BACKGROUND

[0005]

[0002] , Electrochemical sensors have been proven to be useful in the real-time detection of target analytes in diverse media including air, water, process intermediates, and biological fluids. In such sensors, a biological or a chemical analyte recognition element is applied to an electrically conductive element to form a working electrode. Upon interrogation of the sensor by application of a potential, a current is produced. A feature of the current may be used to determine the amount of analyte in the test fluid.

[0006]

[0003] , Electrochemical aptamer-based (EAB) sensors are showing increasing promise in industry and medicine. In an EAB sensor, the working electrode is coated with a plurality of aptamers that specifically recognise a target analyte. A redox reporter (such as methylene blue) is typically covalently linked to the aptamer at its free end. The recognition manifests as a conformational change in the aptamer upon analyte binding. The conformational change in turn alters the accessibility of the redox reporter to the working electrode surface, thereby producing an analyte-induced change in the level of electron transport between the redox reporter and the electrode. The change in speed of electron transport contributes to a change in Faradaic current that is detected by a potentiostat.

[0007]

[0004] , Chronoamperometry is a technique often used in the operation of electrochemical sensors. In this technique, the potential of the working electrode is stepped and the resultant current arising from Faradaic processes at the electrode is measured as a function of time. The Faradaic current is due to electron transfer between the electrode surface and the redox reporter, and is therefore the current component of interest. Since the Faradaic current decays over a relatively long time period, chronoamperometry provides a superior signal- to-noise ratio in comparison to other amperometric techniques.

[0005] , While chronoamperometry reduces noise to some extent, it is still a problem in the art that noise degrades the data output by an electrochemical sensor, and particularly for EAB sensors. This problem may apply to methods other than chronoamperometry. The presence of noise places a restriction on the lower limit of detection for the sensor given that very low signals which are informative as to analyte amounts are masked by the noise. EAB sensors particularly are capable of detecting very low amounts of analyte, however the presence of noise may prevent the sensor from, in practice, achieving its maximum sensitivity.

[0008]

[0006] , It is an aspect of the present invention to provide an improvement in, or an alternative to, prior art electrochemical sensors and the circuits within which they are operable.

[0009]

[0007] , The discussion of documents, acts, materials, devices, articles, and the like, is included in this specification solely for the purpose of providing a context for the present invention. It is not suggested or represented that any or all of these matters formed part of the prior art base or were common general knowledge in the field relevant to the present invention as it existed before the priority date of each claim of this application.

[0010] SUMMARY

[0011]

[0008] , In a first aspect, the present invention provides an electrochemical sensor apparatus comprising: a sensor comprising a working electrode, a counter electrode, and optionally a reference electrode; a power source configured to apply a voltage to the sensor; and a voltage processor comprising one or more components, the voltage processor being configured to reduce a voltage perturbation in the voltage applied to the sensor.

[0012]

[0009] . In one embodiment of the first aspect, the power source comprises a voltage booster circuit.

[0013]

[0010] , In one embodiment of the first aspect, the voltage booster circuit is a DC-DC voltage booster circuit.

[0014] [Oi l], In one embodiment of the first aspect, the electrical potential of the power source is provided by a battery.

[0012] , In one embodiment of the first aspect, the voltage perturbation is an alternating current component, a ripple, a pulse, or noise.

[0015]

[0013] , In one embodiment of the first aspect, the voltage perturbation is capable of negatively affecting the detection of a target analyte by the apparatus.

[0016]

[0014] , In one embodiment of the first aspect, the voltage perturbation is caused by voltage switching in the voltage booster circuit.

[0017]

[0015] , In one embodiment of the first aspect, the voltage processor comprises one or more of a choke, a low pass filter, or a voltage regulator.

[0018]

[0016] , In one embodiment of the first aspect, the choke comprises paired inductors.

[0019]

[0017] , In one embodiment of the first aspect, the low pass filter comprises a capacitor and / or a resistor.

[0020]

[0018] , In one embodiment of the first aspect, the low pass filter is an ElectroMagnetic Compatibility (EMC) filter, an ElectroMagnetic Interference (EMI) filter, or a Radio- Frequency Interference (RFI) filter.

[0021]

[0019] , In one embodiment of the first aspect, the low pass filter is a second order low pass filter.

[0022]

[0020] , In one embodiment of the first aspect, the second order low pass filter is a CL filter or an LC filter.

[0023]

[0021] . In one embodiment of the first aspect, low pass filter is a third order low pass filter.

[0024]

[0022] , In one embodiment of the first aspect, the third order low pass filter is a T-filter or a 7r-filter.

[0025]

[0023] , In one embodiment of the first aspect, the third order low pass filter is a T-filter.

[0026]

[0024] , In one embodiment of the first aspect, the voltage regulator is a linear voltage regulator.

[0027]

[0025] , In one embodiment of the first aspect, the linear voltage regulator is a low dropout voltage regulator.

[0028]

[0026] , In one embodiment of the first aspect, the low pass filter inputs voltage output from the choke.

[0029]

[0027] , In one embodiment of the first aspect, the voltage regulator inputs voltage output from the low pass filter.

[0028] , In one embodiment of the first aspect, the low pass filter inputs voltage output from the choke, and the voltage regulator inputs voltage output from the low pass filter.

[0030]

[0029] , In one embodiment of the first aspect, the choke, the low pass filter, or the voltage regulator, inputs voltage output from the power source.

[0031]

[0030] , In one embodiment of the first aspect, the sensor inputs voltage output from the choke, the T-filter, or the voltage regulator.

[0032]

[0031] , In one embodiment of the first aspect, the sensor inputs voltage output from the voltage regulator.

[0033]

[0032] . In one embodiment of the first aspect, the voltage output from the voltage processor has a noise level sufficiently low so as to at least partially offset the negative effect of the voltage perturbation on the detection of a target analyte by the apparatus.

[0034]

[0033] , In one embodiment of the first aspect, the working electrode and / or counter electrode and / or reference electrode are configured to penetrate the skin of a mammal so as to be contactable to the interstitial fluid, or another bodily fluid thereof.

[0035]

[0034] , In one embodiment of the first aspect, the working electrode and / or counter electrode and / or reference electrode is / are selected from a needle, a microneedle and a wire.

[0036]

[0035] , In one embodiment of the first aspect, the apparatus is configured to be wearable on a surface of a mammal.

[0037]

[0036] , In one embodiment of the first aspect, the apparatus comprises an adhesive surface configured to retain the sensor on the skin of a mammal.

[0038]

[0037] , In one embodiment of the first aspect, the working electrode is coated in a redox- modified aptamer species capable of selective interaction with a target analyte.

[0039]

[0038] , In a second aspect, the present invention provides a method for determining the amount of an analyte in a test fluid, the method comprising contacting the working electrode of the apparatus of any embodiment of the first aspect to the test fluid, applying an interrogating voltage originating in the power source to the working electrode, and measuring a current in the working electrode arising from application of the interrogating voltage.

[0040] BRIEF DESCRIPTION OF THE FIGURES

[0041]

[0039] , FIG. 1A is a graph of voltage versus time of the output of a conventional battery used to power an electrochemical sensor.

[0040] , FIG. IB is a graph of voltage versus time of the output of a conventional voltage booster used to increase the relatively low battery voltage shown in FIG. 1A, to a higher voltage useful to power a sensor. Voltage perturbations are noted.

[0042]

[0041] . FIG. 1 C is a graph of voltage versus time of the output of a voltage processor of the present invention as applied to the voltage booster output shown in FIG IB. It will be noted that the voltage booster increases the relatively low battery voltage shown in FIG. 1 A, to a higher voltage useful to power a sensor with associated electronics such as a microcontroller. A lack of voltage perturbations is noted. In reality, some perturbations would remain although they would be of lesser magnitude than those shown in FIG. IB.

[0043]

[0042] , FIG. 2 is a circuit diagram of a conventional electrochemical sensor apparatus comprising a potentiostat supplying power to the sensor electrodes.

[0044]

[0043] , FIG. 3 is a block diagram of a conventional electrochemical sensor apparatus including a 3.0 V battery as a power source and a voltage booster to increase voltage to an average of 3.2V in accordance with the graph of FIG. IB.

[0045]

[0044] , FIG. 4 is a block diagram of the electrochemical sensor apparatus shown in FIG. 3, although including a voltage processor disposed after the voltage booster. The effect of the voltage processor is to reduce the voltage perturbations as shown in the graph of FIG. IB, to provide power in accordance with the graph of FIG. 1C.

[0046]

[0045] , FIG. 5 is a circuit diagram of a highly preferred voltage processor disposed in an electrical circuit after voltage booster. The voltage processor comprises three components: a choke, a T-filter, and a low dropout voltage regulator.

[0047]

[0046] , Unless otherwise indicated herein, features of the drawings labelled with the same numeral are taken to be the same features, or at least functionally similar features, when used across different drawings.

[0048]

[0047] , The drawings are not prepared to any particular scale or dimension and are not presented as being a completely accurate presentation of the various embodiments.

[0049] DETAILED DESCRIPTION AND PREFERRED EMBODIMENTS

[0050]

[0048] , After considering this description it will be apparent to one skilled in the art how the invention is implemented in various alternative embodiments and alternative applications. However, although various embodiments of the present invention will be described herein, it is understood that these embodiments are presented by way of example only, and not limitation. As such, this description of various alternative embodiments should not be construed to limit the scope or breadth of the present invention. Furthermore, statements of advantages or other aspects apply to specific exemplary embodiments, and not necessarily to all embodiments, or indeed any embodiment covered by the claims.

[0051]

[0049] . Throughout the description and the claims of this specification the word “comprise” and variations of the word, such as “comprising” and “comprises” is not intended to exclude other additives, components, integers, or steps.

[0052]

[0050] , Reference throughout this specification to “one embodiment” or “an embodiment” means that a particular feature, structure, or characteristic described in connection with the embodiment is included in at least one embodiment of the present invention. Thus, appearances of the phrases “in one embodiment” or “in an embodiment” in various places throughout this specification are not necessarily all referring to the same embodiment, but may.

[0053]

[0051] , In seeking to produce a portable or a wearable EAB sensor for use in the real-time monitoring of an analyte in biological fluid of a subject, a prior art sensor (such as that shown in FIG. 2) was powered with two button-cell batteries each of 1.5V. To increase voltage of the battery to a more usable level of 3.2V (particularly to power a microprocessor), a DC-DC voltage booster was incorporated into the circuit. The inventors observed that the voltage booster used to increase battery voltage from 1.5V to a more useful 3.2V resulted in the generation of noise in the EAB sensor apparatus circuit. The noise was found to be sufficient to mask low level current output arising from analyte interaction with the working electrode.

[0054]

[0052] , Without wishing to be limited by theory in any way, it is proposed that the noise arises from rapid switching within the voltage booster. A voltage booster has two operational states: one when the power switch is closed (the switch-on state), and another when the power switch is open (the switch-off state). In relation to the switch-on state, the switch is conducting, and current from the input supply is shunted to ground. Current flows through an inductor, then the switch, and into the ground node. During this time, the inductor magnetic field is increasing. Meanwhile, the load circuitry connected to the booster requires a consistent supply of current which is provided by discharge of an output capacitor which provides the required current during the switch-on state. The inductor is charging, the capacitor is discharging, and a diode blocks current in both directions due to its reverse-biased. In relation to the switch-off state, current flowing through the inductor when the switch was closed, and that inductor current continues to flow when the switch opens, taking the only pathway available: the diode. Voltage boost is achievable because the capacitor can charge to a voltage that is in excess of the supply voltage, so long as current is prevented form flowing back to the source. The boost converter’s diode passes current to the capacitor leading to an increase in voltage. The capacitor cannot discharge back into the source via the inductor because the diode prevents current flow in that direction. The booster circuit rapidly oscillates between the switch-off and switch-on states, and it is proposed that such oscillation is the source of noise in the power output.

[0055]

[0053] , There exists also the potential for switching noise to arise from voltage ripple, which is the result of the capacitor cyclically charging and discharging to cause regular changes in voltage.

[0056]

[0054] , Reference is made to FIG. 1 A showing typical output of a 3.0V battery that may be used to power a portable sensor apparatus. Such voltage may be provided by 2 x 1.5V batteries connected in series. The voltage output has no perturbations, but is of insufficient magnitude to properly power all components of the sensor, such as a microcontroller. A voltage booster of the type discussed supra is therefore introduced after the battery, as shown in FIG. 2.

[0057]

[0055] , Reference is made to FIG. 2 showing a circuit diagram of a prior art electrochemical sensor having an electrode arrangement (10) comprising a working electrode (15), a counter electrode (20), and a reference electrode (25), connected to a potentiostat having a voltage input (Vi). The voltage input is typically provided by way of a high quality mains- powered desktop power source, of the type used in research laboratories. Even with such a power source, noise such as that shown in FIG. IB may be noted, leading to noise- contaminated sensor output.

[0058]

[0056] , Staying with FIG. 2, Vi is an applied potential (the potentiostatic set point) that is referenced to ground. It will be noted in this generic discussion of potentiostat operation the ground may be real or virtual. Vi applies the inverting negative input on the op-amp. The output Vo connects to the counter electrode (20) lead. The reference electrode (25) lead and the working sense lead (15, WEsense) connect to an electrometer (functioning as a true difference amplifier). The output voltage V feedback from the electrometer feeds into a voltmeter V and into the positive non-inverting input of the op-amp. The working electrode (15) drive lead WEr / rzve connects to a resistor R that connects to ground (real or virtual). The voltmeter (E / I), measures the potential across R, and converts it to a current. WE.s s and WEr / rzve are shorted together and connect to the working electrode (15).

[0059]

[0057] , In operation, the user applies voltage Vi to the electrochemical cell. An output voltage (Vo), is output from the op-amp into the electrochemical cell through the counter electrode (20). The counter electrode (20) passes current (ictr) through the test fluid to the reference electrode (25), WEsense, and WEdrive. Inputs to an op-amp have a high impedance, hence no current passes. Since the reference electrode (25) and WE.s s are connected to a high impedance operational amplifier, the only viable path for current flow is through WEdrive.

[0060]

[0058] , The electrometer output voltage is the voltage difference between the reference electrode (25) and WEsense. As the current travels between the counter electrode (20) and WEdrive, the voltage drops across a gradient that is proportional to the bulk solution resistance. This process affects the potential of reference electrode (25) and WEsense. However, the reference electrode is configured to maintain a stable potential, and the potential at reference electrode (25) therefore remains constant. Accordingly any measured changes in the potential originate from WEsense. The voltage difference between reference electrode (25) and WEsense is the output voltage of the electrometer, V feedback. Nfeedback feeds into a voltmeter (V) to measure the difference between the reference electrode (25) and WEsense. It also feeds into the positive non-inverting input of the opamp. Given that the op-amp amplifies the difference between the positive and negative inputs, if fNfeedback s not equal to Vi, the op-amp will either increase or decrease Vountil they equalise.

[0061]

[0059] , Finally, note that in the circuit of FIG. 2, the working electrode current is not directly measured. Rather the voltage is measured across R, which is a known resistor, and based on the voltage measured across it current can be measured by application of Ohm’s Law.

[0062]

[0060] , To summarise, the current at the working electrode is measured by voltmeter (E / I) across R, the potential at the working electrode is measured by the electrometer (V), and the applied potential is determined by the potentiostatic set point. By utilising the op-amp’s feedback mechanism, the potential of the working electrode with respect to the reference electrode can be adjusted. Simultaneously adjusting potential while measuring the current at the working electrode is a feature of techniques used in the interrogation of electrochemical sensors.

[0063]

[0061] , Turning nowto FIG. 3 there is shown a block diagram for an electrochemical sensor apparatus including electrode and potentiostat (40) as shown in FIG. 2, although powered by a battery (28). This apparatus is designed for use as a portable EAB sensor, and particularly a wearable EAB sensor. The battery (28) outputs 3.0V which is insufficient voltage to power on-board microprocessor (45) and other components (50) such as an indicator light, a wireless networking module or a display. Accordingly, a DC-DC voltage booster (30) is used to increase voltage to 3.2V. As just one example, a useful voltage booster may be the MAX17221ENT (Analogue Devices), being an ultra-low quiescent current boost (step-up) DC-DC converters with a 225mA / 0.5A / lA peak inductor current limit. The booster (30) generates noise within the circuit due to switching as discussed above. While the sensor apparatus of FIG. 3 is useful in the context of a wearable sensor, limit of detection problems arise due the contamination of sensor current output with switching noise.

[0064]

[0062] , FIG. 4 shows the circuit of FIG. 3, although with a voltage processor (32) disposed after the booster (30). The function of the voltage processor (32) is to process the output of the booster (30) (having noise in accordance with FIG. IB) so as to be smoothed (in accordance with FIG. 1C) and therefore provide a substantially noise-free current. The voltage processor (32) effectively removes voltages above 3.2V, which at the same time removes the voltage perturbations above 3.2V. The voltage supplied to the potentiostat / electrodes (40) therefore has significantly reduced noise. Accordingly, the current measured (which arises from the interrogation and is informative of the analyte concentration) has less noise contamination.

[0065]

[0063] , As used herein, the term “voltage processor” includes any component or contrivance configured to condition, regularise, or smooth an input voltage.

[0066]

[0064] , As used herein, the term “noise” includes any random or semi-random fluctuations in voltage. Other voltage perturbations may also be problematic, and therefore addressed by the present invention. Such perturbations include any residual alternating current (AC) component in DC output (where the voltage booster is an AC-DC booster), a ripple or a pulse. Some voltage perturbations may not have any connection to a voltage booster, or even any component of the sensor apparatus. For example, perturbations may arise from electromagnetic radiation emitted by proximal electrical or electronic devices such as light fixtures, televisions, automobiles, computers and the like.

[0067]

[0065] , An embodiment of the voltage processor (32) is shown in FIG. 5. This embodiment is highly preferred and has three interconnected components. The first component is a choke (50) which is connected to the booster (30) output. The choke (50) in this embodiment is comprised of paired inductors, arranged as drawn. As just one example of a suitable choke, reference is made to ACM2012-222 -2P-T001 (TDK), being a single line common mode choke. In an alternative embodiment, there is no choke, and instead only a single inductor.

[0068]

[0066] , The second component is a T-filter (50) that is connected to the choke (50). The T- filter (55) is comprised of two shunt inductors and a coupling capacitor, arranged as drawn. Suitable exemplary inductors may be rated at 22 pH and 330 pH, and the capacitor may be rated at 10 pH. The T-filter (55) is configured as a low pass filter, having cut-off in a preferred embodiment of about 27 kHz.

[0069]

[0067] , As an alternative to the T-filter, any other EMF, EMI or RFI filter may be useful. Each of these types of filters, to some extent, mitigate high frequency electromagnetic noise in a power line.

[0070]

[0068] , A jr-filter (being essentially the inverse of a T-filter) may be used. A n-filter consists of an inductor with a ground capacitor on both the input and output. A n-fi Iter presents capacitive terminal characteristics which are desirable when connecting to inductive sources and loads.

[0071]

[0069] , The filter may be any type of first, second or third order filter as applicable to any particular usage context. First order filters consist of one reactive component (such as a capacitor or inductor) and are characterised by a single time constant. They may be used for low-pass and high-pass filtering applications, but preferably low pass in the context of the present invention. Second order filters consist of two reactive components and are characterised by two time constants. They may be used for band-pass and band-stop filtering applications, either of which may be effective in reducing voltage perturbations when appropriately configured. Third order filters have three reactive components and are characterised by three time constants. They can be used for more complex filtering applications, such as band-pass and band-stop filtering with greater selectivity although are also suited to low-pass applications. The second order filter may be an LC filter (an inductor, L, followed by a ground capacitor, C) or a CL filter (ground capacitor, C, as input; and series inductor, L, at output).

[0072]

[0070] , The third component is a LDO (60); being a low dropout voltage regulator that is connected on the input side to the T-filter and on the output side to the potentiostat op-amp.

[0073]

[0071] , An LDO is a type of voltage regulator (and particularly a linear voltage regulator) that is operable at a low potential difference between input and output. In relation to the potential difference between the input and output, an exemplary lower voltage at which the regulator can operate stably is less about or less than IV.

[0074]

[0072] , Operating at a lower potential difference allows for reduce energy losses and suppression of heat generation. Heat generation may be a problem in EAB sensors given that the interaction of analyte with the sensing aptamers is temperature dependant, and calculation of analyte concentration may not take into account any variation in temperature arising from the sensor itself. Energy losses are to be generally avoided where the sensor apparatus is configured as a wearable apparatus (which is typically battery powered) so as to extend battery life.

[0075]

[0073] , In a linear regulator a transistor is placed between Vin and Vo and the minimum potential difference necessary to achieve stable transistor operation is called the dropout voltage. In this way, for both standard linear regulators and LDOs the minimum necessary input voltage is set in order to ensure operation. In this case Vo + dropout voltage is the minimum operating voltage.

[0076]

[0074] , When the input voltage Vin is below the minimum operating voltage the output voltage will not be stable.

[0077]

[0075] , When the voltage difference between the input and output falls below the dropout voltage the transistor cannot maintain stable operation and the output voltage decreases.

[0078]

[0076] , The linear regulator may be series regulator or a shunt regulator. A series regulator, as the name implies, is placed in series between a power source and a load whereas a shunt regulator is placed in parallel between a power source and a load. Either a series or a shunt regulator may be useful in the context of the present invention dependent upon the application.

[0079]

[0077] , Linear regulators do not rely on any switching, and accordingly no new switching noise will be introduced by this component.

[0080]

[0078] , The functions of any two components of the voltage processor may overlap to some extent, or the functions may be entirely discrete.

[0081]

[0079] , In some embodiments, the voltage processor is selected from a choke, a low pass filter or a voltage regulator.

[0082]

[0080] , In some embodiments, the voltage processor comprises a choke and a low pass filter, or a choke and a voltage regulator, or a low pass filter and a voltage regulator.

[0083]

[0081] , In some embodiments, the voltage processor comprises a choke, a low pass filter, and a voltage regulator.

[0084]

[0082] , In some embodiments, the voltage processor is only one, two, three or four elementary electronic components selected independently from: an inductor, a capacitor, a transistor, and a resistor.

[0085]

[0083] . The ability to detect target analyte to low limits by way of an electrochemical sensor is of significant practical advantage in many applications, but particularly clinical applications. As an example of such application, and electrochemical sensor may be an EAB sensor having a working electrode coated in a redox-modified aptamer that is specific for the cardiac marker troponin. The electrodes of the sensor are each a microneedle capable of penetrating the skin of a subject (such as a human) and contacting the underlying interstitial fluid (ISF). Upon interrogation the working electrode of the sensor detects the concentration of troponin in the ISF. Such a sensor may be used in the diagnosis of a cardiac condition. Troponin may be present at very low levels, and accordingly the electrochemical sensor should have a correspondingly low limit of detection for that analyte. Failure to detect a low level of troponin may result in a missed diagnosis of acute myocardial infarction.

[0086]

[0084] , The invention may be useful in other applications such as in sensors for the determination of clinically relevant analytes in a pathology laboratory.

[0085] , The invention may have applicability also in non-clinical applications such as the monitoring of target analyte concentrations in environmental settings (such as waste water), and industrial settings (such as process intermediates).

[0087]

[0086] . The present invention may be embodied in the form of a fully functional EAB sensor, having a working electrode coated with analyte-specific DNA, RNA or XNA. Gold is often used as the probe surface for the working electrode. The aptamer has an associated redoxactive species which acts as a reporter. The redox reporter is often methylene blue. Upon target (e.g., drug) binding, the aptamer undergoes a conformational change, bringing the redox reporter more proximal to the working electrode surface. This increase in proximity increases electron transfer from the redox reporter to the electrode. The increase in speed of electron transfer contributes to a change in Faradaic current that is detected by a potentiostat.

[0088]

[0087] , Aptamers are small (usually from 20 to 60 nucleotides) single-stranded RNA, DNA or XNA oligonucleotides able to bind a target drug with high affinity and specificity. Aptamers may be considered as nucleotide analogues of antibodies, but aptamer production is an in vitro cell-free process that is significantly easier and cheaper than the production of antibodies by cell culture or in vivo methods.

[0089]

[0088] , Aptamers are usually selected from combinatorial library having a vast number (up to 1015) of different oligonucleotides. While RNA aptamers provide a significantly greater structural diversity compared to DNA aptamers, their application is complicated by stability issues in the presence of Rnases, high temperature, and unfavourable pH.

[0090]

[0089] , Selection of an aptamer that is selective for a given drug may be facilitated by a process known as SELEX (systematic evolution of ligands by exponential enrichment). The process may be considered as two alternating stages. In the first stage, the library oligonucleotides are amplified by a polymerase chain reaction (PCR) to the desired concentration. For the selection of RNA aptamers, the single-chained oligoribonucleotides are generated by in vitro transcription of double-stranded DNA with T7 RNA-polymerase. For DNA aptamers, a pool of single-stranded oligodeoxyribonucleotides is generated by strand separation of double-stranded PCR products. In the second stage, the products of amplification are incubated with target drug and oligonucleotides which bind the drug are used in the next SELEX round.

[0090] , Separation of oligonucleotides with higher affinity for target drug and removal of unbound oligonucleotides are achieved through intense competition for binding sites. The selection pressure rises with every SELEX round. Maximum enrichment of the oligonucleotide pool with aptamers with the strongest affinity for the target molecule is usually achieved after 5 to 15 rounds.

[0091]

[0091] , EAB sensors are typically incorporated into a circuit having a reference electrode. The reference electrode is the site of a known chemical reaction that has a known redox potential. For example, a reference electrode based on the silver-silver chloride (Ag|AgCl) redox pair has a fixed and known potential forming the point against which the redox potential of the working electrode is measured. Also typically included in the circuit is a counter electrode which functions as a cathode or an anode to the working electrode. Because the applied voltage bias does not pass through the reference electrode (due to an impedance of the potentiostat), any potential generated is attributed to the working electrode. Current is measured as potential of the interrogating electrode versus the stable potential of the reference electrode. The difference in potential produces the current in the circuit thereby generating an output signal. The signal quantifies target binding depending on electron transfer that is ideally stoichiometrically proportional to target binding.

[0092]

[0092] , EAB sensors are particularly suitable for use as a wearable apparatus, allowing measurements to be performed whilst the subject is undergoing normal activities and / or over a prolonged period of time. The wearable apparatus may be a collar, a bracelet or other suitable jewellery piece, a watch, a garment, a strap, an adhesive, or a patch. A person skilled in the art would appreciate that means may be provided to assist adhering and / or securing the wearable apparatus, when in use, to a subject, e.g., micro-anchors, or the like.

[0093]

[0093] , The wearable apparatus may comprise a housing structure comprising one or more other components, such as electronics processing unit. The electronics processing unit is configured to be in direct or indirect electrical communication with at least one electrode, and generally will include any one or more of a power source, a data processing unit, an analogue front-end, and a wireless transmitter.

[0094]

[0094] , The housing structure may be configured to encase, at least partially, the apparatus, where the electrodes (such as microneedles) are exposed from a plane of the housing structure. The electrodes may be protected by a protective cover, which may be removed to expose the protruding electrodes before use.

[0095]

[0095] , The apparatus may further comprise means for monitoring temperature or pH of the biological fluid where validity of an output is dependent thereon, or where adjustment to operation or output is possible.

[0096]

[0096] , The housing structure may be configured to encase and be coupled to the apparatus by any appropriate mechanism. For example, electromagnetic coupling, mechanical coupling, adhesive coupling, magnetic coupling, or the like. In some embodiments, the coupling mechanism enables the apparatus and the housing structure to be attached and detached, which would enable the housing structure and its other components to be reusable, while the apparatus can be discarded and replaced with another apparatus as necessary.

[0097]

[0097] , The wearable apparatus may further comprise a computer program product executable as a software application, resident on a mobile communication device in communication with the electronics processing unit, wherein the computer program product is able to control one or more of (i) detection of electrochemical measurements conducted at the electrode-based platform, (ii) data analysis, (iii) data transmission, (iv) apparatus configuration, and (v) apparatus power management. Examples of suitable mobile communication devices include, but are not limited to, smartphones, smartwatches, tablets, smartglasses, laptops or other personal computers.

[0098]

[0098] , In some embodiments, the apparatus itself comprises a processor with program instructions configured to drive onboard functions such as voltammetry, and transmitting output to a remote device via a wireless module, such as a Bluetooth™ module.

[0099]

[0099] , As will be understood, the present invention may be deployed in part or in whole through one or more microprocessors that execute computer software, program codes, and / or instructions on a processor. A microprocessor may be any kind of computational or processing device capable of executing program instructions, codes, binary instructions, and the like.

[0100]

[0100] , Any microprocessor may access a storage medium (such as electronic memory) through an interface that may store methods, codes, and instructions as described herein and elsewhere. The storage medium associated with the processor for storing methods, programs, codes, program instructions or other type of instructions capable of being executed.

[0101]

[0101] , The computer software, program codes, and / or instructions may be stored and / or accessed on computer readable media that may include: computer components, devices, and recording media that retain digital data used for computing for some interval of time; semiconductor storage known as random access memory (RAM); mass storage typically for more permanent storage, such as non-volatile memory such as read only memory (ROM).

[0102]

[0102] , The methods described herein may transform physical and / or or intangible items from one state to another. The methods and systems described herein may also transform data representing physical and / or intangible items from one state to another.

[0103]

[0103] , Software products may be created using a structured programming language such as C, an object oriented programming language such as C++, or any other high-level or low-level programming language (including assembly languages, hardware description languages, and database programming languages and technologies) that may be stored, compiled or interpreted to run on a microprocessor, as well as heterogeneous combinations of processors, processor architectures, or combinations of different hardware and software, or any other machine capable of executing program instructions.

[0104]

[0104] , Thus, in one aspect, any method may be embodied in computer executable code that, when executing on one or more microprocessors, performs the steps thereof. In another aspect, the methods may be embodied in systems that perform the steps thereof, and may be distributed across devices in a number of ways, or all of the functionality may be integrated into a dedicated, standalone device or other hardware. In another aspect, the means for performing the steps associated with the processes described above may include any of the hardware and / or software described above. All such permutations and combinations are intended to fall within the scope of the present disclosure.

[0105]

[0105] , The invention may be embodied in program instruction set executable on one or more microprocessors. Such instructions set may include any one or more of the following instruction types.

[0106]

[0106] , Data handling and memory operations, which may include an instruction to set a register to a fixed constant value, or copy data from a memory location to a register, or vice-versa, to store the contents of a register, result of a computation, or to retrieve stored data to perform a computation on it later, or to read and write data from hardware devices.

[0107]

[0107] , Arithmetic and logic operations, which may include an instruction to add, subtract, multiply, or divide the values of two registers, placing the result in a register, possibly setting one or more condition codes in a status register, to perform bitwise operations, e.g., taking the conjunction and disjunction of corresponding bits in a pair of registers, taking the negation of each bit in a register, or to compare two values in registers (for example, to determine if one is less, or if they are equal).

[0108]

[0108] , Control flow operations, which may include an instruction to branch to another location in the program and execute instructions there, conditionally branch to another location if a certain condition holds, indirectly branch to another location, or call another block of code, while saving the location of the next instruction as a point to return to.

[0109]

[0109] , Coprocessor instructions, which may include an instruction to load / store data to and from a coprocessor, or exchanging with CPU registers, or perform coprocessor operations.

[0110]

[0110] , A processor of a computer of the present system may include “complex” instructions in their instruction set. A single “complex” instruction does something that may take many instructions on other computers. Such instructions are typified by instructions that take multiple steps, control multiple functional units, or otherwise appear on a larger scale than the bulk of simple instructions implemented by the given processor. Some examples of “complex” instructions include: saving many registers on the stack at once, moving large blocks of memory, complicated integer, and floating-point arithmetic (sine, cosine, square root, etc.), SIMD instructions, a single instruction performing an operation on many values in parallel, performing an atomic test-and-set instruction or other read-modify-write atomic instruction, and instructions that perform ALU operations with an operand from memory rather than a register.

[0111]

[0111] , An instruction may be defined according to its parts. According to more traditional architectures, an instruction includes an opcode that specifies the operation to perform, such as add contents of memory to register — and zero or more operand specifiers, which may specify registers, memory locations, or literal data. The operand specifiers may have addressing modes determining their meaning or may be in fixed fields. In very long instruction word (VLIW) architectures, which include many microcode architectures, multiple simultaneous opcodes and operands are specified in a single instruction.

[0112]

[0112] , Some types of instruction sets do not have an opcode field (such as Transport

[0113] Triggered Architectures (TTA) or the Forth virtual machine), only operand(s). Other unusual “0-operand” instruction sets lack any operand specifier fields, such as some stack machines including NOSC.

[0114]

[0113] , Conditional instructions often have a predicate field — several bits that encode the specific condition to cause the operation to be performed rather than not performed. For example, a conditional branch instruction is executed, and the branch taken, if the condition is true, so that execution proceeds to a different part of the program, and not executed, and the branch not taken, if the condition is false, so that execution continues sequentially. Some instruction sets also have conditional moves, so that the move is executed, and the data stored in the target location, if the condition is true, and not executed, and the target location not modified, if the condition is false. Similarly, IBM z / Architecture has a conditional store. Some instruction sets include a predicate field in every instruction; this is called branch predication.

[0115]

[0114] , The instructions constituting a program are rarely specified using their internal, numeric form (machine code); they may be specified using an assembly language or, more typically, may be generated from programming languages by compilers.

[0116]

[0115] , While the invention is described mainly by reference to a microneedle electrode, the invention is equally applicable to an electrode of any form and dimension.

[0117]

[0116] , Similarly, the invention is described mainly by reference to the use of an electrochemical sensor to sense an analyte in the ISF of a mammal. The invention may be operable with other test fluids such as blood, saliva, urine, potable water, waste water, industrial process intermediates, industrial products, foods, and many others.

[0118]

[0117] , Accordingly, the spirit and scope of the present invention is not to be limited by the foregoing examples, but is to be understood in the broadest sense allowable by law.

Claims

AMENDED CLAIMS received by the International Bureau on 08 August 2025 (08.08.2025)CLAIMS:

1. An electrochemical sensor apparatus comprising: a sensor comprising a working electrode, a counter electrode, and optionally a reference electrode; a power source configured to apply a voltage to the sensor; and a voltage processor comprising a choke, a low pass filter, and a voltage regulator, the voltage processor being configured to reduce a voltage perturbation in the voltage applied to the sensor.

2. The apparatus of claim 2, wherein the power source comprises a voltage booster circuit that is a DC-DC voltage booster circuit, or another voltage booster circuit.

3. The apparatus of claim 1 or claim 2, wherein the electrical potential of the power source is provided by a battery.

4. The apparatus of any one of claims 1 to 3, wherein the voltage perturbation is an alternating current component, a ripple, a pulse, or noise.

5. The apparatus of claim 4, wherein the voltage perturbation is capable of negatively affecting the detection of a target analyte by the apparatus.

6. The apparatus of any one of claims 2 to 5, wherein the voltage perturbation is caused by voltage switching in the voltage booster circuit.

7. The apparatus of any one of claims 1 to 6, wherein the choke, the low pass filter, and the voltage regulator are electrically connected in that order.

8. The apparatus of claim 7, wherein the choke comprises paired inductors or the low pass filter comprises a capacitor and / or a resistor.

9. The apparatus of claim 7 or claim 8, wherein the low pass filter is an ElectroMagnetic Compatibility (EMC) filter, an ElectroMagnetic Interference (EMI) filter, or a Radio-Frequency Interference (RFI) filter.

10. The apparatus of any one of claims 7 to 9, wherein the low pass filter is CL filter or an LC filters, or another second order low pass filter.

11. The apparatus of any one of claims 7 to 10, wherein the low pass filter is a third order low pass filter that is a T-filter or a a-filtcr or another third order low pass filter.

12. The apparatus of any one of claims 7 to 11, wherein the voltage regulator is a linear voltage regulator.

13. The apparatus of claim 12, wherein the linear voltage regulator is a low dropout voltage regulator.

14. The apparatus of any one of claims 7 to 13, wherein the low pass filter inputs voltage output from the choke.

15. The apparatus of any one of claims 7 to 14, wherein the voltage regulator inputs voltage output from the low pass filter.

16. The apparatus of any one of claims 7 to 15, wherein the low pass filter inputs voltage output from the choke, and the voltage regulator inputs voltage output from the low pass filter.

17. The apparatus of any one of claims 7 to 16, wherein the choke, the low pass filter, or the voltage regulator, inputs voltage output from the power source.

18. The apparatus of any one of claims 7 to 17 wherein the sensor inputs voltage output from the choke, the T-filter, or the voltage regulator.

19. The apparatus of any one of claims 1 to 18, wherein a voltage output from the voltage processor has a noise level sufficiently low so as to at least partially offset the negative effect of the voltage perturbation on the detection of a target analyte by the apparatus.

20. The apparatus of any one of claims 1 to 19, wherein the working electrode is coated in a redox-modified aptamer species capable of selective interaction with a target analyte.

Images