This invention relates to a sensor and in particular to a sensor for the detection of biologically important species.
Modern healthcare relies extensively on a range of chemical and biochemical analytical tests on tissue samples and a variety of body fluids to enable early recognition, diagnosis and management of disease. Accordingly, there is a significant market for in vitro diagnostics, including sensors.
Chemical sensors have found widespread use in numerous applications. Biosensors are examples of such sensors. In order to create a sensor for a specific analyte these sensors typically contain a chemical recognition element coupled to a means of signal transduction. The presence of the analyte of interest causes a measurable change in a property of the transduction material. A wide range of transduction modalities to convert the physico-chemical response to the analyte in the test medium into the measurement signal have been developed. Examples include amperometric, potentiometric, conductimetric, optical, gravimetric, surface-acoustic waves (SAW), thermal or capacitive principles.
To date chemically selective membranes and biomolecules, such as proteins (e.g. enzymes, antigens, antibodies), nucleic acids (DNA or RNA) or even whole microorganisms have been used as recognition elements. More recently, synthetic materials, such as synthetic receptors and molecularly imprinted polymers (MIPs) have been employed to replace the traditionally used recognition elements.
One of the most challenging issues in sensor manufacture relates to the reliable attachment or integration of the receptors with the transducer. In particular for multi-use devices, the receptor needs to be stable, robust and maintain its selectivity. It has to stay attached to the transducer over the complete period of use without degradation or leaching. Moreover, the receptor should continue to function despite the attachment/integration with the transducer surface; similarly the transducer should remain functional despite the attachment or integration of the receptor.
In addition, the use of chemical sensors is often affected by interferents, which affect the sensor response and lead to erroneous readings. The detection of the analyte of interest when bound to the receptor is also problematic.
Accordingly, the present invention provides a sensor for detecting an analyte in the presence of at least one interferent in a fluid sample comprising
a substrate,
a channel disposed on the substrate,
at least one transducer disposed in the channel, and
a filter for selectively absorbing the analyte in preference to the at least one interferent, wherein the filter is arranged in the channel remotely from the at least one transducer and wherein the channel defines a flow path for the fluid sample such that the fluid sample contacts a first transducer to generate a first signal, followed by the filter, followed by the first transducer or a second transducer to generate a second signal.
That is, the sensor comprises a selective filter which removes the analyte from the fluid sample allowing a measure to be taken before and after removal of the analyte. The filter is positioned to hold the analyte remotely from the transducer(s). This allows the analyte to be measured without having to bind the analyte selectively to a transducer which has significant practical difficulties.
The present invention will now be described with reference to the accompanying drawings, in which:
The present invention relates to a novel arrangement for chemical sensing and to a chemical sensor system. It provides a straight-forward means for detecting the presence of one or more analytes of interest in the sample being investigated and/or for measuring their concentration(s). One particular application area for these devices and systems relates to the detection and measurement of medical drugs, markers or medically relevant substances which are indicative of the health, status or treatment of a patient. The sensor or sensor system can, for example, be used to analyse the current state of health of the patient and/or direct treatment of a medical condition suffered by the patient. In this context, the term “patient” refers to both humans and animals. Furthermore, the term “sample” refers to a substance or mixture of substances being investigated and includes, in particular, but not limited to, bodily fluids, such as blood, urine, interstitial fluids and cerebro-spinal fluids.
In order to resolve any issues associated with the attachment/integration of receptors with a transducer, the sensor of the present invention has a filter 4 which does not need to be bound to a transducer.
By subtracting the signal generated by the second transducer 3 (located behind the filter) from the signal generated by the first transducer 2, a signal characteristic of the amount or concentration of the analyte of interest in the sample is obtained. The amount of the analyte(s) of interest contained in the sample can be calculated using, for example, appropriate sensor calibration. Rather than determining directly the presence of the analyte(s) of interest in the sample, the sensor 1 of the present invention determines the presence and/or concentration of analyte(s) of interest present in the sample by detecting the absence of the analyte removed from the sample by passage through the filter.
It is important that the fluid sample contacts the first transducer 2 to generate a first signal, followed by the filter 4, followed by the second transducer 3 to generate a second signal. In order to achieve this order of events, the channel 6 in the sensor 1 defines a flow path for the fluid sample which allows the fluid sample to contact the transducers 2,3 and filter 4 in this order. The direction of flow is indicated in
In the sensor 1 of the present invention, the filter is arranged in the channel remotely from the at least one transducer. By remote is meant that the filter is sufficiently remote that any analyte present in the filter is not detected by the at least one 15 ‘ transducer. If the analyte were detected by the transducer, the second signal could not be reliably subtracted from the first signal. This is a fundamentally different approach to known sensors, such as that disclosed in WO 2005/075995, where the transducer incorporates a material for selective binding of the analyte, but the bound analyte is detected by the transducer. In the sensor of the present invention, it is the absence of the analyte which is detected.
A key advantage of the sensor 1 of the present invention is that the receptor for the analyte(s) of interest is not integrated on top of the transducer surface, but can be disposed as a filter 4 in a convenient format remote from the transducers. Preferably the filter 4 is located between the first and second transducers 2,3. It therefore removes a number of manufacturing difficulties, is quicker and more cost-effective to produce and provides more design freedom. Moreover, this approach avoids any issues associated with the integration of receptor and transducer on the sensor performance.
The channel 6 is shown in
A wide range of materials for the filter 4 can be used. These include chemically selective membranes (for example, polymer membranes containing ionophores which are known in the art and are commercially available from suppliers such as Sigma Aldrich) and biomolecules, such as proteins (e.g. enzymes, antigens, antibodies), nucleic acids (DNA or RNA), microorganisms or synthetic materials, such as synthetic receptors and molecularly imprinted polymers.
The synthetic polymer may be any synthetic polymer provided the polymer is capable of selectively binding an analyte (i.e. it functions as a receptor). The selective binding may be a result of functional groups on the polymer which interact with a specific analyte. The synthetic polymer preferably comprises one or more functionalised monomers and one or more cross-linkers. A preferred polymer is a molecularly imprinted polymer.
Molecularly imprinted polymers (MIPs) are essentially artificial macromolecular receptors prepared by molecular imprinting of synthetic polymers. MIPs are prepared by polymerising functional monomers or copolymerising functional and cross-linking monomers in the presence of an imprint molecule which acts as a molecular template. The functional monomers initially form a complex with the imprint molecule and, following polymerisation, their functional groups are held in position by the highly cross-linked polymeric structure. In this way, a molecular memory is introduced into the polymer which is then capable of binding the imprint molecule. The imprinting of small organic molecules is now well established in the art.
The functional monomer should be capable of binding to the imprint molecule, via functional groups on the functional monomer. Binding may be via a covalent bond or by intramolecular forces, such as a hydrogen bond or van der Waals forces. A suitable functional group may be, for example, a carboxylic acid in a (meth)acrylic acid ester, although the nature of the functional group will depend on the nature of the imprint molecule. The monomer must, of course, be polymerisable and able to react with a cross-linker when present. Suitable monomers include, but are not limited to, acrylic monomers, such as (meth)acrylic acid, (meth)acrylic acid esters, (meth)acrylamide, (meth)acrylonitrile, 2-hydroxyethylmethacrylate (HEMA), N,N,N-triethylaminoethyl(meth)acrylate, trifluoromethylacrylic acid, acrylamide, N,N′-methylenebisacrylamide, acrylonitrile, 2-acrylamido-2-methyl-1-propanesulfonic acid acrolein, ethylene glycol dimethacrylate, imidazole-4-acrylic acid ethyl ester, imidazole-4-acrylic acid, 2-(diethylamino)ethyl methacrylate; vinyl and allyl monomers, such as 2- and 4-vinylpyridine, m-and p-divinylbenzene, styrene, aminostyrene 1-vinylimidazole, allylamine; urethanes; phenols; boronates, such as aminophenyl boronate; amines, such as phenylene diamine, phenylene diamine-co-aniline; organosiloxane monomers; carbonate esters, such as methylenesuccinic acid; sulfonic acid; or mixtures (i.e. co-polymers) thereof. See M. Komiyama et al.
Molecular Imprinting: From Fundamentals to Applications, Wiley-VCH Verlag GmbH & Co KGaA, Weinheim (2003), G. Wulff Angew. Chem. Int. Ed. Engl. 34, 1812 (1995), and S. Subrahmanyam et al. Biosensors & Bioelectronics 16, 631 (2001).
The cross-linker may be included to fix the template-binding sites firmly in the desired structure as, well as to influence the porosity of the MIP. The cross-linker must be capable of reacting with the functional monomers to cross link the polymer chains and the cross-linker should preferably be of similar reactivity to the monomer. Suitable cross-linkers include, but are not limited to, ethylene glycol dimethacrylate (EDMA), glycerol dimethacrylate (GDMA), trimethylacrylate (TRIM), divinylbenzene (DVB) (which is particularly suitable for cross linking acrylate- and vinyl-containing functional monomers), methylenebisacrylamide and piperazinebisacrylamide (which are particularly suitable for cross linking acylamides), phenylene diamine (which is particularly suitable for cross linking amines such as aniline and aminophenyl boronate), dibromobutane, epichlorohydrin, trimethylolpropane trimethacrylate and N,N′-methylenebisacrylamide.
The mole ratio of functional monomer to cross-linker is preferably from 1:1 to 1:15. Mixtures of monomers and cross-linkers may also be used.
The functional monomer and/or the cross-linker may act as a solvent for the polymerisation reaction or an additional solvent may be added. Suitable solvents are known in the art and include DMSO (dimethyl sulfoxide), formic acid, acetic acid, DMF (dimethylformamide), methanol, acetonitrile, dichloromethane, chloroform, THF (tetrahydrofuran), toluene and cyclohexane. Mixtures of these solvents may also be used to obtain the desired solvation and porogenic properties.
The polymer preferably has a molecular weight from 1 to 100,000 kDa, more preferably 10 to 10,000 kDa and most preferably 10 to 5,000 kDa.
See WO 2005/075995, WO 2006/120381 and WO 2006/120382 for further details regarding MIPs and other synthetic polymers which may be used with the sensor 1 of the present invention.
In the case of a sensor for the anaesthetic drug propofol, one preferred class of materials to be used in the filter are molecular imprinted polymers (MIP). For example, a non-covalent MIP capable of binding propofol and composed of methacrylic acid (MAA) as the functional monomer and ethylenedimethylacrylic acid (EDMA) as the cross-linker. See WO 02/00737 and WO 2006/120381 for further details.
The filter 4 is preferably positioned such that the whole of the fluid sample passes through the filter 4, as shown in
In the embodiment shown in
In another embodiment of the invention,
Although the sensors shown in
The amperometric sensor can be of two electrode or three-electrode design. The electrodes may be made of a number of materials, including silver, gold, platinum, carbon, stainless steel. Other materials are known to those skilled in the art. The amperometric sensor may also be functionalised through the deposition of enzymes (for example, glucose oxidase, lactate oxidase, or other suitable chemicals), membranes (for example, comprising silicone, HEMA, PVC, Nafion, polyurethane) in order to detect preferentially the analyte of interest and remove the effects of interferents. One example to functionalise a sensor for the measurement of glucose in this manner is described in U.S. Pat. No. 6,440,296.
While the preferred embodiment of this invention has been described with particular reference to amperometric sensors, a large number of transduction principles can be used to realise the invention. These include potentiometric (in particular, ion selective field effect transistors, ISFETs, and chemically selective field effect transistor, CHEMFETs), conductimetric, optical, gravimetric, surface-acoustic waves, resonant, capacitive or thermal principles. The sensors may be functionalised to preferentially detect the analyte of interest, to detect a group of analytes of interest or to reduce the effect of interferents through the deposition of chemically selective membranes (for example, polymer membranes containing ionophores) and biomolecules, such as proteins (e.g. enzymes, antigens, antibodies), nucleic acids (DNA or RNA), microorganisms or synthetic materials, such as synthetic receptors and molecularly imprinted polymers. Many methods to achieve this are known to those skilled in the art.
Rather than employing transducers of the same type, one embodiment of the sensor uses one transducer type upstream of the filter, for example an amperometric transducer, while a different transducer type is employed downstream of the filter, for example, a potentiometric sensor. The responses from both sensors are then converted to provide an indication of the total concentration of the analyte of interest and interferents at the location of the transducer using, for example, appropriate sensor calibrations. The difference in the concentration measurements of the two transducers is related to the absorption of the analyte of interest on the filter and therefore to the concentration of the analyte of interest in the sample.
In another preferred embodiment of the invention, the sensor is a micromachined device. The transducers employed in the chemical sensor can be realised on the same die or different dies. One particular example of a silicon-based microsensor chip with multiple transducers is shown in
In addition to filtering out the effect of interferents, the differential approach utilised in the invention is also able to remove any disturbance or effect, which affects both transducers. It is therefore able to compensate, for example, for temperature-related effects, changes in the supply voltage for the transducers, flow speed variations etc.
Following the measurement, the filter 4 may be regenerated, for example by passing an appropriate fluid through the sensor chamber, applying temperature changes or electrical potential changes in order to remove any material absorbed on the filter. Alternatively, the filter may be designed in such a way that it possesses enough capacity to enable multiple uses or continuous operation without regeneration. This latter approach is of particular advantage for sensor for continuous use or in an application, where regeneration of the filter is not possible.
In another embodiment of the invention, the adsorption of the analyte(s) of interest will release another agent into the sample, which is subsequently detected by a transducer which is capable of detecting the agent. In this case, the increase in the sensor signal associated with the release of the agent in the fluid is measured.
While most embodiments in this invention have been described with reference to a construction employing a single continuous substrate 5, another embodiment of the invention can be realised by mounting each transducer on its own substrate and connecting them in series. The filter is then integrated into the fluid connection between the two substrates.
In another embodiment of the invention, the sensor is adapted to detect a first analyte and a second or further analyte. In this case, the sensor contains one or more filters for selectively absorbing the first, second or further analyte in preference to the at least one interferent. Where a number of filters are used, one filter may specifically absorb one analyte, while a second filter employed in series to the first filter may specifically absorb a second analyte of interest, etc.
Within the area of clinical and medical diagnostics, the sensor disclosed herein is particularly useful for the analysis of blood with respect to the presence and concentration of markers or substances indicative of the health or status of a patient or drugs used in the treatment of a patient's condition. In particular, sensors based on the invention can be applied to the following areas: measurement of anaesthetic agents, in particular those administered intravenously, such as propofol (propofol can be administered in a variety of forms, for example as an emulsion or in an aqueous forms); detection and measurement of antibiotic agents, e.g. penicillin and vancomycin; detection and measurement of heart markers, such as troponins, ischaemia-modified albumin etc.; and measurement of electrolytes and gases dissolved in bodily fluids, such as blood and urine.
The sensor of the present invention may be adapted to detect any of the above analytes. The adaptation is apparent from the breadth at which the signal is detected, e.g. the voltage range selected for a potentiometric transducer.
The sensor of the present invention is typically incorporated into a sampling system and a signal processing unit. Accordingly, the present invention also provides a sampling apparatus comprising a housing coupled to a sampling port and incorporating the sensor as described herein and a signal processing unit in electronic communication with the sensor. An example of such a system is shown in
The sensor 1 is connected to a local display and signal processing unit 12 which may be connected to a patient monitoring device 13. The sensor 1 is also connected to the housing 8 electronically using techniques known in the art.
In addition to the system described above, the sensor may be employed in a range of other sensing systems, known to those skilled in the art. For example, rather than being directly connected to the patient, a sample may be taken from the patient and transported to and injected into an analyser, into which the sensor is integrated, for sample analysis.
In addition to providing detection and measurements of markers, substances or drugs, the sensor of the present invention provides feedback for the treatment of the patient based on the results of the analysis made. This feedback may be provided either directly to the user or it may be part of a closed-loop control system including the device administering the treatment to the patient. One particular example is a sensor for an anaesthetic agent, such as propofol, which measures the concentration of the anaesthetic agent in one or more bodily fluids or body compartments, e.g. blood or blood plasma, and based on these measurements directs, either directly or the user, the subsequent delivery of the anaesthetic agent, e.g. by controlling the rate of delivery to the patient via a syringe pump.
The sensor may also be used with systems which monitor other parameters which characterise the health of a patient, monitor particular markers indicating disease states or direct the patient's treatment, e.g. blood gases, pH, temperature etc.
The present invention further provides a method of detecting an analyte in the presence of at least one interferent in a fluid sample comprising
providing a fluid sample potentially containing the analyte,
contacting the fluid sample with the sensor as described herein,
obtaining the first and second signals, and
comparing the first and second signals to provide an indication of the amount of the analyte present in the sample.
Number | Date | Country | Kind |
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0524580.8 | Dec 2005 | GB | national |
Filing Document | Filing Date | Country | Kind | 371c Date |
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PCT/GB06/04496 | 12/1/2006 | WO | 00 | 7/25/2008 |