Patent ID: 12215374

DETAILED DESCRIPTION OF THE EMBODIMENTS

In a first aspect of the invention there is provided an organic thin film transistor based sensor for detecting the presence of an analyte in a liquid sample, the sensor including:a porous wicking layer having a first surface and a second surface, wherein the first surface is configured to receive a liquid sample;an enzyme disposed on or within the porous layer, the enzyme for facilitating the generation of a charge carrier from an analyte;a polymer gating layer in contact with the second surface of the porous layer, the polymer gating layer configured to enable an ohmic conductor to connect to the polymer gating layer for applying a gate voltage to the polymer gating layer, the polymer gating layer being conductive to the charge carrier; andan organic semiconducting layer configured to be connected to a source electrode and a drain electrode.

An exemplary organic thin film transistor based sensor100in accordance with one embodiment of the invention is illustrated inFIG.1, which provides a conceptual representation of the structure of the sensor100. Sensor100includes a drain electrode102and source electrode104disposed on the surface of a substrate106. The sensor includes a three-layered film structure that includes: a porous wicking layer108, a polymer gating layer120, and an organic semiconducting layer122. The organic semiconducting layer122covers a portion of the drain and source electrodes, with the organic semiconducting layer108in contact with, and extending between the drain electrode102and the source electrode104. The polymer gating layer120is disposed on a surface of the organic semiconducting layer122. An ohmic conductor132is in contact with polymer gating layer120to enable a gate voltage to be applied to the polymer gating layer120. A porous wicking layer108is disposed on the surface of the polymer gating layer120. The surface of the porous wicking layer108is exposed to receive a fluid sample. An enzyme is disposed within and/or on the surface of the porous wicking layer108. The porous wicking layer100is not located between gate and drain electrodes. The porous wicking layer100is not configured to enable an ohmic conductor to connect to the porous wicking layer.

The inventors have found that incorporation of a porous wicking layer including an enzyme into an organic thin film transistor is advantageous. This is for two primary reasons. Firstly, on depositing a liquid sample onto the surface of the porous wicking layer, the porous wicking layer exhibits a wicking effect which improves incorporation of a liquid sample into the device, this wicking effect has the potential to ensure delivery of a standard volume of liquid sample to within the device. Secondly, the wicking effect results in excellent wetting of the internal porous structure of the porous wicking layer which results in a high degree of exposure of an analyte within the liquid sample to the enzyme retained within the porous wicking layer. This is beneficial as it results in efficient generation of charge carriers within the device, and consequently the rapid detection of the analyte if present within the liquid sample.

The skilled person will appreciate that the porous wicking layer108may be formed from a range of different materials, including: porous ceramics, porous metals, porous protein fibres, or porous polymers. However, generally porous polymers are preferred for ease of sensor fabrication. Suitable polymers include, but are not limited to: polyacrylonitrile, polysulfone, polyethersulfone, polystyrene, polybutadiene, polyisoprene, polyimides, polyamides, and fluoropolymers. Typically the porous wicking layer is formed from a material that is not the same as the material that the polymer gating layer is formed from. Where the polymer gating layer is formed from Nafion, the porous wicking layer is not formed from Nafion. Preferably the porous wicking layer is not formed from Nafion.

Ideally the porous wicking layer108is formed from a material that has a low contact angle with the liquid of the liquid sample to ensure that the liquid sample is rapidly taken into the porous wicking layer108and that there is good wetting of pore surfaces of the porous wicking layer108.

Preferably the porous wicking layer takes the form of a layer that enables the rapid formation of a droplet, preferably an aqueous droplet, more preferably a biological fluid droplet, more preferably a saliva droplet, such as a neat saliva droplet or a processed or diluted saliva droplet, on the porous wicking layer that has a low contact angle with the porous wicking layer.

Preferably the porous wicking layer is formed from a layer having a high degree of wettability by an aqueous droplet, more preferably a biological fluid droplet, more preferably a saliva droplet, more preferably a neat saliva droplet or a processed or diluted saliva droplet.

Notwithstanding the above, it is preferred that the porous wicking layer108is a porous poly(acrylonitrile) (PAN) layer. PAN has physical properties that make it particularly useful as the porous wicking layer108. PAN is thermally stable with a glass transition temperature (Tg) of ˜95° C. and melting temperature (Tm)>300° C. Given this, PAN exhibits good thermal stability, which is desirable in a sensor as this means that during typical use, the Tgis unlikely to be exceeded. If the Tgis exceeded, the porous structure may collapse. PAN also has high strength, a high modulus of elasticity, and a low density. Furthermore, from a fabrication point of view, PAN is soluble in dimethyl sulfoxide (DMSO) and can be used to prepare porous polymer membranes via the phase inversion technique-immersion precipitation.

In a preferred embodiment, the porous wicking layer has a sufficiently low surface energy to enable a body fluid, preferably saliva, preferably undiluted or otherwise unprocessed saliva, or processed saliva to rapidly wet and penetrate the porous wicking layer.

As discussed above, the porous wicking layer108includes an enzyme. The enzyme may be disposed on a surface of the porous wicking layer108and/or disposed throughout the porous wicking layer108, such that the enzyme is exposed on pore surfaces within the porous wicking layer108.

The enzyme is selected to facilitate generation of charge carriers when an analyte contacts the device. The charge carriers are typically electrons, anions, or cations (e.g. hydrogen ions/protons). The generation of the charge carriers may be further facilitated by the presence of an electric field. As will be described, these generated charge carriers can then contribute to electric current through the device. It will be recognised that a range of enzymes could be used for any one particular analyte. Further given the diversity of enzymes available, the device, by following the disclosure herein can be adapted or developed for detection of a range of analytes.

A particularly preferred class of enzyme is an oxidoreductase. An oxidoreductase for use in the device may act on any one of the following donor groups: the CH—OH group of donors (alcohol oxidoreductases), the aldehyde or oxo group of donors, the CH—CH group of donors (CH—CH oxidoreductases), the CH—NH2group of donors (amino acid oxidoreductases, monoamine oxidase), CH—NH group of donors, NADH or NADPH, other nitrogenous compounds as donors, a sulfur group of donors, a heme group of donors, diphenols and related substances as donors, peroxide as an acceptor (peroxidases), hydrogen as donors, single donors with incorporation of molecular oxygen (oxygenases), paired donors with incorporation of molecular oxygen, superoxide radicals as acceptors, CH or CH2groups, iron-sulfur proteins as donors, reduced flavodoxin as a donor, phosphorus or arsenic in donors, X—H and Y—H to form an X—Y bond, or oxidoreductases that oxidize metal ions.

The polymer gating layer120is disposed between the porous wicking layer108and the organic semiconducting layer122. The function of polymer gating layer120is to facilitate transport of charge carriers generated in the porous wicking layer108from an analyte on application of a gate voltage. A range of different polymers may be used to form the polymer gating layer120depending on the nature of the analyte, enzyme, and/or charge carrier. In a preferred form, the charge carriers are protons (such as hydrogen ions) and the polymer gating layer120is proton conductive. Preferably the polymer layer has a conductivity to protons that is greater than a conductivity to protons of the organic semiconductor layer122. The conductivity may be due to permeability of the polymer gating layer120to charge carriers, such as where conduction occurs via migration of the charge carriers; alternatively, conduction may occur via another mechanism, such as the Grotthuss mechanism. Where a charge carrier is a proton, or is an electron, the polymer gating layer may be proton conductive or electron conductive.

In preferred forms of the invention, the polymer gating layer120comprises, consists, or consists essentially of a sulfonated tetrafluoroethylene-based fluoropolymer-copolymer, for example a copolymer comprising a tetrafluoroethylene backbone and perfluoroalkyl ether groups terminated with sulfonate groups. More preferably, the sulfonated tetrafluoroethylene-based fluoropolymer-copolymer is a copolymer of tetrafluoroethylene and perfluoro-3,6-dioxa-4-methyl-7-octene-sulfonic acid. An example of a suitable polymer is tetrafluoroethylene-perfluoro-3,6-dioxa-4-methyl-7-octenesulfonic acid copolymer (commonly referred to as Nafion).

In preferred forms of the invention the polymer gating layer has a thickness in the range of from about 50 nm to about 5 μm. It is preferred that the thickness is from about 200 to about 800 nm.

The organic semiconducting layer122is configured to enable flow of electrical current between the source electrode and the drain electrode as a result of the generation of these charge carriers.

The organic semiconducting layer122includes one or more organic compounds. Any organic compound having semiconducting properties is suitable for use. However, it is preferred that the one or more organic compounds are selected from the group consisting of: polyacetylenes, porphyrins, phthalocyanins, fullerenes, polyparaphenylenes, polyphenylenevinylenes, polyfluorenes, polythiophenes, polypyrroles, polypyridines, polycarbazoles, polypyridinevinylenes, polyarylvinylenes, poly (p-phenylmethylvinylenes), including derivatives and co-polymers thereof, and further including combinations thereof. More preferably, the one or more organic compounds are selected from the group consisting of: poly(9,9-dioctylfluorene-2,7-diyl-co-bis-N, N-(4-butylphenyl)-bis-N, N-phenyl-1,4-phenylenediamine), poly(9,9-dioctylfluorene-2,7-diyl-co-benzothiadiazole), poly(3-hexylthiophene), (6,6)-phenyl-C61-butyric acid methyl ester, poly(2-methoxy-5-(2′-ethyl-hexyloxy)-1,4-phenylene vinylene), and combinations thereof. Most preferably, the semiconducting layer includes, consists of, or consists essentially of P3HT.

In preferred forms of the invention, the organic semiconducting layer has a layer thickness of from 20 nm up to 500 nm. Preferably, the layer thickness is up to 350 nm. More preferably, the layer thickness is up to 200 nm. Most preferably, the layer thickness is up to 150 nm. Alternatively, or additionally, the layer thickness is from 40 nm. More preferably the layer thickness is from 60 nm. Most preferably, the layer thickness is from 70 nm.

The use of the sensor100will now be described below in relation to a preferred embodiment in which the sensor100is for detecting the presence of glucose in a saliva sample.

In use, a gate voltage VGand a drain voltage VD are applied to the sensor100(the voltages being with respect to the source electrode104, as shown inFIG.1), and a liquid sample comprising glucose, for example a bodily fluid such as saliva, is contacted with the porous wicking layer108. The liquid sample is wicked into the internal porous structure of the porous wicking layer, where glucose in the liquid sample comes into contact with glucose oxidase enzyme (GOX) that is present within the porous wicking layer. The glucose is degraded via an enzymatic reaction with GOX thereby producing H2O2. The gate voltage VGand drain voltage VDprovide a sufficiently strong electric field to liberate H+from H2O2, but not strong enough to cause electrolysis of water, as electrolysis of water may lead to a decrease in the signal-to-noise ratio of the sensor. Typically, the gate voltage and drain voltage applied are between about 0 V and −2 V, e.g. about −1 V.

The H+ions are conducted from the porous wicking layer108though the polymer gating layer120(e.g. Nafion) to the organic semiconducting layer. This results in doping of the semiconductor, and consequentially, current between the drain and the source electrodes. Without wishing to be bound by theory, the inventors are of the view that the gate potential controls the doping and de-doping of the semiconducting compound(s) via ion migration from the site of ion generation to the active channel in the organic semiconductor. Thus, the increase in H+ions results in an increase in drain current, such that a relationship is established between the amount of glucose present in the sample and the magnitude of the drain current. The drain current is then measured which provides an indication of the presence and concentration of glucose within the saliva sample.

The gate voltage VGand drain voltage VDprovide a sufficiently strong electric field to liberate H+from H2O2, but not enough to cause electrolysis of water, as electrolysis of water may lead to a decrease in the signal-to-noise ratio of the sensor. In at least one embodiment, the gate voltage and drain voltage applied are between about 0 V and −2 V, e.g. about −1 V.

In an alternative form of the invention, as illustrated inFIG.2, the sensor200is similar to sensor100ofFIG.1, but further comprises a dielectric layer234between and in contact with polymer gating layer220and organic semiconductor layer222.

The dielectric layer234comprises, consists of, or consists essentially of an organic dielectric material. Preferably, the organic dielectric material has a conductivity to protons that is greater than the conductivity of organic semiconductor layer222. Preferably the organic dielectric material is a hygroscopic insulator, such as for example polyvinyl phenols. More specifically, the dielectric layer may comprise, consist of, or consist essentially of, poly(4-vinylphenol).

Alternative dielectric materials that may be used in the devices will be readily apparent to those skilled in the art. Non-limiting examples include polyimide and poly(methyl methacrylate (PMMA). In alternative embodiments the dielectric layer may comprise a doped dielectric material, for example lithium perchlorate doped poly(4-vinylpyridine).

EXAMPLE

Fabrication of the Device

Pre-patterned ITO-on-glass substrates (15 Ω□−1ITO, Xin Yan Technology) were used as the substrate and electrodes of the devices. Poly-3-hexylthiophene (P3HT) (MW ˜20 000) was dissolved in CHCl3(Sigma-Aldrich) at various concentrations and sonicated for ˜1 hour or until the material was entirely dissolved. Poly-4-vinylphenol (PVP) (Sigma-Aldrich) was dissolved in ethanol (Sigma-Aldrich) at a concentration of 80 mg mL−1and sonicated for ˜1 hour or until the material was entirely dissolved. Nafion solution (5% by weight in lower aliphatic alcohols and water, Sigma-Aldrich) was used as received.

The pre-patterned ITO-on-glass substrates were cleaned with methanol and purified water. P3HT solution in CHCl3was spin-coated onto the substrates at 2000 rpm for 60 seconds. P3HT solutions of 5 mg mL−1, 10 mg mL−1, 15 mg mL−1, 20 mg mL−1, and 40 mg mL−1were prepared, with average thicknesses of films spun from these concentrations of P3HT were 22 nm, 36 nm, 74 nm, 108 nm and 390 nm respectively. The P3HT layer was patterned and then left to dry for 15 minutes at 40° C. Nafion solution was first spin-coated at 500 rpm for 120 seconds.

While the devices exemplified herein do not include a dielectric layer, a dielectric layer may be included. A preferred dielelectric layer is a PVP layer. For devices including a PVP layer, PVP solution can be spun on top of the P3HT layer at 2000 rpm for 60 seconds to form a film of thickness ˜400 nm, which can then patterned and dried. For PVP-containing devices, Nafion can then be drop-cast above the source-drain channel area and connected to the ITO gate pad of the substrate before being dried for approximately 30 minutes.

Porous polymer membranes can be fabricated using the phase inversion (immersion precipitation) technique. Firstly, a viscous solution of polyacrylonitrile (PAN) polymer in DMSO solvent is coated on a flat, rigid substrate, such as glass, to form a uniform ‘wet’ film. The ‘wet’ film can be coated by either spin coating, drop-casting or with the use of a printing technique such as slot-die coating. The ‘wet’ film is not a solid, dry film, it is a composite film, comprising a polymer network with liquid solvent molecules existing in the voids between polymer chains. The ‘wet’ film upon the rigid substrate is then submerged in water (the non-solvent), the porous polymer membrane forming as the DMSO solvent diffuses out of the polymer network into the surrounding water bath and the water diffuses into the polymer network. It is this solvent exchange with non-solvent that induces precipitation of the polymer. The selection of solvent and non-solvent is highly dependent on the chosen polymer, the solvent needs to be capable of dissolving the polymer whereas the non-solvent is one in which the polymer is insoluble. The solvent and non-solvent combination should be miscible, otherwise the solvent will not diffuse into the non-solvent and the porous polymer membrane will not form, instead remaining as a ‘wet’ solvent-containing film. The fabrication is a two-step process, it is during the second step that the ‘wet’ film transforms into a solid film. Once the solid film has been formed, it is processed into individual layers of the desired size and thickness and applied to the device as a porous wicking layer.

To incorporate GOX into the wicking layer, the GOX is cast on top of the wicking layer. The porosity of the wicking layer promotes the dispersion of the GOX throughout the wicking layer.

Testing of the Device

The porosity of PAN membranes prepared via the phase inversion technique were investigated using scanning electron microscopy (SEM) (seeFIG.3) on a Zeiss ZP FESEM operating at an accelerating voltage of 2 kV. DMSO was used as the solvent and water as the non-solvent in the membrane preparation process. SEM revealed PAN membrane pore size to be in the 200-800 nm size range.

To investigate the potential for wicking of PAN membranes, the contact angle of water on PAN compared to water on Nafion was measured.FIG.4presents photographs of water droplets on (a) PAN and (b) Nafion from which the contact angle was extracted. The contact angle of water on PAN was measured to be ˜35°, this increased to ˜70° on Nafion. The decreased contact angle of water on PAN indicates that PAN is a more hydrophilic surface than Nafion. This reduced contact angle on PAN is beneficial for a wicking effect, an excellent result for initial trials of this material.

X-ray photoelectron spectroscopy (XPS) was performed to probe any potential redistribution of Nafion throughout the PAN porous membranes following analyte addition. XPS spectra were collected by illuminating the samples with a non-monochromatic X-ray source (Omnivac) using Al Kα (1486.6 eV) radiation, and the photoemission collected by an SES2002 analyser (Scienta). The F 1s peak was utilised as a marker for Nafion as Nafion contains fluorine atoms attached to the carbon atoms of the polymer backbone whereas PAN only contains the elements carbon, nitrogen and hydrogen. Structures for (a) PAN and (b) Nafion are provided below:

An XPS survey scan (FIG.5(a)) of a pristine Nafion film identified the F 1s peak at a binding energy of 691 eV. XPS region scans were then performed of a pristine PAN membrane, a Nafion/PAN bilayer and a Nafion/PAN bilayer following exposure to analyte solution. The pristine PAN membrane had no F 1s peak (FIG.5(b)). The Nafion/PAN bilayer XPS spectrum contains a peak for F 1s, the Nafion/PAN bilayer following exposure to analyte solution XPS spectrum also contains a peak for F 1s, and this peak is quite invariant compared to the Nafion/PAN bilayer indicating that the analyte solution does not cause solvation and redistribution of the underlying Nafion film. This is a positive result for the biosensors indicating that the Nafion film is quite robust.

Glucose sensors were fabricated with the architecture depicted inFIG.6. Sensors with PAN and without PAN were fabricated and the GOX layer was inkjet printed with a Dimatix DMP 2831 inkjet printer with a 10 pL nozzle printing head (FIG.7).

The sensors were exposed to glucose analyte solutions and the difference in response of sensors with and without PAN porous membranes was analysed.FIG.7(b)presents representative plots of a sensor with PAN and a sensor without PAN following exposure to a 10 mM glucose solution. Here we observe an improvement in the drain current (ID) response time as well as the maximum current when we utilise porous PAN membranes. The rate of ID increase changes from 0.04 μA/s without PAN to 0.93 μA/s with PAN, and we observe a more than three-fold improvement in total ID for the PAN-containing sensor. The superior response time suggests that glucose is oxidised at a faster rate and/or the hydrogen peroxide is able to travel to the Nafion layer more quickly.

It will be understood that the invention disclosed and defined in this specification extends to all alternative combinations of two or more of the individual features mentioned or evident from the text or drawings. All of these different combinations constitute various alternative aspects of the invention.