Patent Description:
Paper-based rapid tests such as lateral flow tests provide a cheap and easy-to-use way of detecting specific target substances, which for example is widely used in medical diagnostics, e.g. for pregnancy tests and antigen tests like COVID-<NUM> antigen tests. These tests often rely on optical detection methods such as visual inspection or measurements of reflectivity or fluorescence intensity, which either do not allow for obtaining quantitative results or require complex and expensive equipment.

Electrochemical detection methods such as voltammetry, on the other hand, enable quantitative measurements with simple and inexpensive equipment, but are challenging to implement in paper-based rapid tests. The electrodes required for performing such measurements may for example be placed on the paper substrate. In this case, however, only a small fraction of the sample fluid flowing within the paper substrate comes in contact with the electrode, which limits the measurement sensitivity that can be achieved. <NPL> discloses the formation of a carbon nanofiber electrode by laser-induced carbonization of polymer nanofibers on different substrates such as indium tin oxide (ITO), aluminum foil, glass slides and wax paper. <NPL> discloses the preparation of Ni nanoparticles embedded in three-dimensional carbon nanofibers using electrospinning followed by laser carbonization under ambient conditions. <NPL> discloses a laser process for generating carbon nanofiber nonwovens from polyacrylonitrile by producing carbon nanofabrics via electrospinning followed by infrared laser-induced carbonization.

It is thus an object of the invention to provide means for performing quantitative electrochemical measurements with high sensitivity on liquid samples in a cheap and easy-to-use test setup.

This object is met by a method of forming a carbon nanofiber electrode according to claim <NUM>, a method of manufacturing a sensor for performing an electrochemical measurement on a liquid sample according to claim <NUM>, and a sensor for performing an electrochemical measurement on a liquid sample according to claim <NUM>. Embodiments of the present invention are detailed in the dependent claims.

The method of forming a carbon nanofiber electrode according to the invention comprises providing an electrically insulating porous substrate and forming a mat of electrically insulating nanofibers on the substrate by electrospinning. The electrically insulating nanofibers comprise an organic polymer. An electrode comprising electrically conductive carbon nanofibers is formed in the mat of electrically insulating nanofibers by laser-induced carbonization of the electrically insulating nanofibers.

The electrically insulating porous substrate may for example comprise or consist of an electrically insulating foam and/or electrically insulating fibers, e.g. electrically insulating microfibers. A plurality of pores or channels may extend through the substrate, wherein an average diameter of the pores may for example be between <NUM> and <NUM>, in one example between <NUM> and <NUM>. The diameter of the pores may vary between individual pores, wherein the diameters of the pores in the substrate may for example be distributed within a range between <NUM> and <NUM>. The plurality of pores may form an interconnected network of channels extending through the substrate. An electrical conductivity of the substrate may for example be below <NUM>-<NUM> (Ω · m)-<NUM>, in some examples below <NUM>-<NUM> (Ω · m)-<NUM>, in one example below <NUM>-<NUM> (Ω · m)-<NUM>. The substrate may for example have an electrical conductivity between <NUM>-<NUM> (Ω · m)-<NUM> and <NUM>-<NUM> (Ω · m)-<NUM>, in some examples between <NUM> · <NUM>-<NUM> (Ω · m)-<NUM> and <NUM> · <NUM>-<NUM> (Ω · m)-<NUM>. Preferably, the substrate comprises or is formed of paper, i.e. cellulose fibers. Additionally or alternatively, the substrate may e.g. comprise or consist of nitrocellulose. The substrate may in particular comprise or be formed of filter paper. The substrate may for example comprise or consist of filter paper having a particle retention capacity of less than <NUM>, in some examples less than <NUM>, in one example less than <NUM>. The particle retention capacity of a filter paper may for example be defined as the diameter of the smallest particles that are filtered by the filter paper with an efficiency equal to or larger than a predefined efficiency value, i.e. the diameter of the smallest particles of which the filter paper retains at least a fraction of the particles that is equal to the predefined efficiency value, which may e.g. be <NUM>%, in some examples <NUM>%, in one example <NUM>%. A thickness of the substrate may for example be between <NUM> and <NUM>, in some examples between <NUM> and <NUM>.

In a preferred embodiment, the method further comprises performing a hydrophilic surface treatment on the substrate prior to forming the mat of electrically insulating nanofibers thereon. The hydrophilic surface treatment may make a surface of the substrate less hydrophobic/more hydrophilic, e.g. to increase adhesion of the electrically insulating nanofibers to the substrate. The hydrophilic surface treatment may for example comprise a chemical and/or physical modification of the surface of the substrate, e.g. by creating hydrophilic groups on the surface and/or by adsorption and/or absorption of one or more hydrophilic substances on the surface. In other embodiments, a hydrophilic substrate may be provided and no hydrophilic surface treatment may be performed on the substrate prior to forming the mat of electrically insulating nanofibers thereon.

Performing the hydrophilic surface treatment may in particular comprise performing a plasma treatment on the substrate. This may create oxygen-containing radicals in and/or on the substrate, which may improve adhesion between the mat of electrically insulating nanofibers and the substrate. The plasma treatment may for example comprise exposing the substrate to a plasma for a duration between <NUM> and <NUM>, preferably between <NUM> and <NUM>. A power used for generating the plasma may for example be set to between <NUM> Wand <NUM> W, in some examples between <NUM> W and <NUM> W. The plasma may for example be an O<NUM> plasma, an Ar plasma, and/or an N<NUM> plasma.

Additionally or alternatively, the hydrophilic surface treatment may for example also comprise exposing the substrate to ultraviolet light, performing a chemical surface treatment on the substrate and/or providing electric charges on the substrate. The method may for example comprise exposing the substrate to atomic oxygen, e.g. by performing a UV/ozone treatment. A chemical surface treatment may modify the surface of the substrate through chemical reactions to increase its hydrophilicity, e.g. by exposing the substrate to an acidic solution, for example to induce aminolysis and/or hydrolysis, and/or by covalent grafting of a hydrophilic substance such as a hydrophilic polymer on the substrate, for example by exposing the substrate to a solution comprising the hydrophilic substance. Electric charges may for example be provided on the substrate by adsorption and/or absorption of a charged species, e.g. by exposing the substrate to a solution comprising a polyelectrolyte, and/or by charging the substrate electrostatically, e.g. by inducing a corona discharge above the substrate (as for example also used in xerography, e.g. for laser printing).

The mat of electrically insulating nanofibers is formed on the substrate by electrospinning. The substrate may for example be provided on an electrically conductive collector plate, e.g. a metallic collector plate. A spinning solution comprising the organic polymer may be supplied to a nozzle or orifice arranged above the collector plate. The organic polymer may for example be dissolved in a non-polar solvent or preferably in a polar solvent. The solvent may for example be selected from the group consisting of methylene chloride, ethylene chloride, chloroform, tetrachloroethane, tetrahydrofurane (THF), dioxane, acetophenone, cyclohexanone, m-cresol, g-butyrolactone, dimethylformamide (DMF), dimethylacetamide (DMAC), and N-methylpyrrolidone (NMP). An electric voltage may be applied between the nozzle and the collector plate and the spinning solution may be ejected from the nozzle towards the collector plate. The spinning solution may solidify and may form an interconnected web of electrically insulating nanofibers comprising the organic polymer on the substrate arranged on the collector plate. In the following, the electrically insulating nanofibers in the mat may also be referred to as polymer nanofibers. Properties of the polymer nanofibers such as their thickness and density as well as the thickness of the resulting mat of polymer nanofibers may be controlled by adjusting parameters of the electrospinning process such as a composition of the spinning solution, a molecular weight of the polymer, a viscosity of the spinning solution, a conductivity of the spinning solution, a flow rate of the spinning solution through the nozzle, a duration of the electrospinning process, a distance between the nozzle and the collector plate, the voltage applied between the nozzle and the collector plate, a temperature at which the electrospinning is performed, and/or a humidity at which the electrospinning is performed. Preferably, the substrate is rotated while forming the polymer nanofiber mat thereon to improve the uniformity of the mat. The substrate may for example be rotated continuously while performing the electrospinning or may be rotated one or more times in discrete steps, e.g. by <NUM>° or <NUM>°. In some embodiments, the method may comprise drying the mat of polymer nanofibers, e.g. to remove liquid remaining in the polymer nanofiber mat.

In a preferred embodiment, the polymer nanofibers are formed by electrospinning of a spinning solution comprising the organic polymer and a metal-containing substance. The metal-containing substance may for example be a metal acetylacetonate, in particular tris(acetylacetonato)iron(III), (Fe(acac)<NUM>). Providing a metal-containing substance in the spinning solution may increase a thermal conductivity of the polymer nanofiber mat, which may be advantageous to prevent burning of the polymer nanofiber mat and/or of the substrate during the laser-induced carbonization. A concentration of the metal-containing substance in the spinning solution may for example be between <NUM>/ml and <NUM>/ml, in some examples between <NUM>/ml and <NUM>/ml, preferably between <NUM>/ml and <NUM>/ml. In one example, the concentration of the metal-containing substance in the spinning solution is between <NUM>/ml and <NUM>/ml, e.g. <NUM>/ml. A power and/or a scanning rate of a laser beam used for the laser-induced carbonization may be adapted to the concentration of the metal-containing substance, e.g. by using a lower laser power and/or a faster scanning rate for higher concentrations of the metal-containing substance and vice-versa.

The organic polymer may be a polymer containing carbon atoms in its backbone and may for example be an organic thermoplastic. The organic polymer may in particular be selected from the group consisting of polyimide, polyacrylonitrile, poly(amic acid), poly(p-xylenetetrahydrothiophenium chloride), polybenzimidazole, poly(vinyl alcohol), and poly(vinylidene fluoride). Preferably, the organic polymer is polyimide. A concentration of the organic polymer in the spinning solution may for example be between <NUM>/ml and <NUM>/ml, preferably between <NUM>/ml and <NUM>/ml, in one example <NUM>/ml. In some examples, the electrically insulating nanofibers may comprise multiple organic polymers, for example a mixture of two or more of the aforementioned polymers.

Parameters of the electrospinning process, in particular the duration of the electrospinning process, may be chosen such that a thickness of the polymer nanofiber mat is between <NUM> and <NUM>, preferably between <NUM> and <NUM>, in one example between <NUM> and <NUM>. The duration of the electrospinning process may for example be between <NUM> and <NUM>, in some examples between <NUM> and <NUM> per cm<NUM> of surface area of a collecting area that is to be coated with the mat of polymer nanofibers. A diameter of a laser beam used for the laser-induced carbonization may be adapted to the thickness of the polymer nanofiber mat, e.g. by using a laser beam with a smaller diameter for thinner polymer nanofiber mats and vice-versa. An average diameter of the polymer nanofibers in the mat may for example be between <NUM> and <NUM>, in some examples between <NUM> and <NUM>. The diameter of the polymer nanofibers may vary between individual nanofibers, wherein the diameters of the polymer nanofibers in the mat may for example be distributed within a range between <NUM> and <NUM>. The polymer nanofibers may in the mat form an interconnected web or mesh of fibers. A plurality of pores or channels may extend through the polymer nanofiber mat such that a liquid may enter or penetrate the mat by capillary transport. An average diameter of the pores may for example be on the order of <NUM> to <NUM>, in some examples between <NUM> and <NUM>.

One or more electrodes comprising electrically conductive carbon nanofibers are formed in the mat of polymer nanofibers by laser-induced carbonization. This may for example comprise scanning a laser beam, in particular a focused laser beam, across the mat of polymer nanofibers, e.g. to selectively heat portions of the mat of polymer nanofibers in which the one or more electrodes are to be formed. By exposing the polymer nanofibers to the laser beam, the polymer nanofibers may be carbonized at least in part, thereby forming electrically conductive carbon nanofibers. This allows for forming conductive electrodes of arbitrary shapes in the mat of polymer nanofibers. An intrinsic electrical conductivity of the carbon nanofiber electrodes may for example be larger than <NUM> (Ω · m)-<NUM>, in some examples larger than <NUM> (Ω · m)-<NUM>, in one example larger than <NUM> (Ω · m)-<NUM>. A power of the laser beam may for example be between <NUM> W and <NUM> W, in some examples between <NUM> W and <NUM> W. A scanning rate of the laser beam may for example be between <NUM>/s and <NUM>/s, in some examples between <NUM>/s and <NUM>/s. The laser-induced carbonization may for example be carried out in an inert gas atmosphere or preferably in ambient atmosphere, i.e. in air.

Each of the carbon nanofibers may for example comprise or consist of graphite and may e.g. comprise a large number of graphene sheets that are stacked on top of each other to form the respective carbon nanofiber. The carbon nanofibers may on average be thicker and/or less uniform than the polymer nanofibers, e.g. because two or more polymer nanofibers may melt and merge at least in part during the laser-induced carbonization. In other examples, the carbon nanofibers may have a similar shape and/or similar physical dimensions as the polymer nanofibers. In contrast to carbon nanotubes, carbon nanofibers may comprise a plurality of defects, e.g. sites at edges of the graphene sheets, thus providing many active sites for transferring an electric charge to or from an electroactive substance, e.g. an electroactive substance in a sample fluid surrounding the carbon nanofibers. A plurality of pores or channels may extend through the electrode such that a liquid may enter or penetrate the electrode by capillary transport, wherein the pores may on average have a larger diameter than the pores in non-carbonized portions of the polymer nanofiber mat in some examples. A sample fluid may thus surround the carbon nanofibers in the electrode, thereby greatly increasing the electroactive surface area. This makes carbon nanofiber electrodes a powerful tool for electrochemical measurements, see e.g. <NPL>.

The formation of a carbon nanofiber electrode by laser-induced carbonization of polymer nanofibers on a substrate consisting of indium tin oxide (ITO) has for example already been reported in <NPL> and <NPL>. While an electrically conductive substrate such as ITO may increase the uniformity of the electric field during electrospinning and may thus improve the uniformity of the resulting polymer nanofiber mat, its electrical conductivity prevents the formation of multiple electrodes thereon. Separating the polymer nanofiber mat from the conductive substrate may be challenging due to the high risk of damaging the polymer nanofiber mat and/or the electrodes formed therein. Furthermore, when carbon nanofiber electrodes are formed in a polymer nanofiber mat by laser-induced carbonization after separation from the conductive substrate, the polymer nanofiber mat may be burned due to its poor thermal conductivity, non-uniformity, and/or folds or wrinkles formed therein.

The present inventors have observed that using an electrically insulating porous substrate such as filter paper allows for forming polymer nanofiber mats with high uniformity by electrospinning as well as for forming carbon nanofiber electrodes by laser-induced carbonization without burning the polymer nanofiber mat and/or the substrate. Moreover, the electrically insulating porous substrate facilitates releasing the polymer nanofiber mat from a collector plate used for electrospinning. The polymer nanofiber mat can also be peeled off the substrate easily, yielding a free-standing nanofiber mat with one or more carbon nanofiber electrodes formed therein.

In a preferred embodiment, the method further comprises performing a hydrophilic surface treatment on the polymer nanofiber mat. The hydrophilic surface treatment can for example be performed after forming the carbon nanofiber electrode. The polymer nanofibers and/or the carbon nanofibers may initially be hydrophobic and the hydrophilic surface treatment may be used for making the polymer nanofibers and/or the carbon nanofibers less hydrophobic or even hydrophilic, e.g. if the carbon nanofiber electrode is to be used for measurements on an aqueous solution. Similar to the hydrophilic surface treatment on the substrate described above, the hydrophilic surface treatment on the polymer nanofiber mat may for example comprise a chemical and/or physical modification of surfaces of the polymer nanofibers and/or of the carbon nanofibers, e.g. by creating hydrophilic groups on the surfaces and/or by adsorption and/or absorption of one or more hydrophilic substances on the surfaces. In other embodiments, no hydrophilic surface treatment may be performed on the polymer nanofiber mat, e.g. if the carbon nanofiber electrodes are to be used for measurements on a solution comprising alcohol and/or an amphiphilic substance such as a detergent.

Performing the hydrophilic surface treatment on the polymer nanofiber mat may in particular comprise performing a plasma treatment on the polymer nanofiber mat. The plasma treatment may for example comprise exposing the substrate to a plasma for a duration between <NUM> and <NUM>, preferably between <NUM> and <NUM>, e.g. <NUM>. The plasma may for example be an O<NUM> plasma, an Ar plasma, and/or a N<NUM> plasma. Additionally or alternatively, the method may also comprise activating the polymer nanofibers and/or the carbon nanofibers by other means. The hydrophilic surface treatment on the polymer nanofiber mat may for example also comprise an exposure to ultraviolet light, a chemical surface treatment and/or providing electric charges on the polymer nanofibers and/or on the carbon nanofibers, e.g. as described above for the hydrophilic surface treatment on the substrate. In some embodiments, the hydrophilic surface treatment on the polymer nanofiber mat may for example comprise exposing the polymer nanofiber mat to atomic oxygen, e.g. by performing a UV/ozone treatment, and/or by exposing the polymer nanofiber mat to an acid, e.g. sulfuric acid (H<NUM>SO<NUM>).

In some embodiments, the method further comprises functionalizing the carbon nanofiber electrode. This may for example comprise adding one or more functional groups to the carbon nanofibers, in particular to an outer surface of the carbon nanofibers. The one or more functional groups may for example comprise one or more electrochemically active groups and/or one or more functional groups that are configured to bind to or immobilize a specific target substance, e.g. a biomolecule such as a specific protein or a specific antibody or antigen. The carbon nanofiber electrode may in particular be functionalized by adsorbing, absorbing and/or embedding a catalyst, in particular nanoparticles and/or enzymes, in the carbon nanofiber electrode. The nanoparticles may e.g. be metallic nanoparticles, which may for example comprise or consist of silver, gold, platinum, palladium, nickel, copper, iron or a combination thereof. Additionally or alternatively, the nanoparticles may e.g. comprise or consist of titanium dioxide, silicon dioxide, molybdenum disulfide and/or a composite material. The enzymes may for example be or comprise glucose oxidase, alcohol oxidase, alcohol dehydrogenase, acetylcholine esterase, tyrosinase, laccase, horseradish peroxidase or a combination thereof. In some embodiments, the carbon nanofiber electrode may be functionalized by adsorbing, absorbing and/or embedding quantum dots, e.g. nanoparticles comprising a quantum dot. Additionally or alternatively, the carbon nanofiber electrode may for example be functionalized by adsorbing, absorbing and/or embedding an ion exchanger, e.g. a cation exchanger or an anion exchanger, in the carbon nanofiber electrode. In one example, a sulfonated tetrafluoroethylene-based fluoropolymer-copolymer such as Nafion is used as a cation exchanger. Other examples of suitable ion exchangers include poly(styrene sulphonate), poly(vinyl sulphate), deprotonated poly(acrylic acid) and/or poly(-L-lysine), Tosflex, poly(vinylpyridine) and/or cellulose acetate, zeolites, Amberlite IR <NUM> or combinations thereof. In some embodiments, the carbon nanofiber electrode may additionally or alternatively also by functionalized by providing a redox mediator such as methylene blue, by molecular imprinting, by providing a conducting polymer such as polyaniline, polypyrrole and/or poly(<NUM>,<NUM>-ethylenedioxythiophene) polystyrene sulfonate, and/or by providing a functional biocompatible polymer such as polydopamine and/or hydrogel.

A substance for functionalizing the carbon nanofiber electrode, e.g. metallic nanoparticles and/or a metal-containing substance, may for example be provided in the spinning solution. In one example, the spinning solution may comprise nickel acetylacetonate, e.g. in addition to or instead of Fe(acac)<NUM>, in order to form nickel nanoparticles embedded in the carbon nanofiber electrode. Additionally or alternatively, a substance for functionalizing the carbon nanofiber electrode may for example be provided after forming the carbon nanofiber electrode, e.g. by exposing the carbon nanofiber electrode to a solution comprising the substance to allow for adsorption and/or absorption of the substance on a surface of the carbon nanofibers. In one example, the carbon nanofiber electrode is exposed to a solution comprising a sulfonated tetrafluoroethylene-based fluoropolymer-copolymer such as Nafion, wherein a concentration of the sulfonated tetrafluoroethylene-based fluoropolymer-copolymer in the solution may for example be between <NUM>/ml and <NUM>/ml, preferably between <NUM>/ml and <NUM>/ml.

The method may further comprise forming one or more hydrophobic barriers in the mat of electrically insulating nanofibers. The one or more hydrophobic barriers may for example define a channel extending through one or more carbon nanofiber electrodes. The one or more hydrophobic barriers may be configured to prevent water from leaking out of the channel into other portions of the polymer nanofiber mat, thereby confining the water to the channel along one or more directions. In one example, a hydrophobic barrier is formed that extends along a closed loop in the polymer nanofiber mat, thereby enclosing a channel in polymer nanofiber mat. The hydrophobic diameter may for example extend along the edge of a rectangle, e.g. to define a channel having a rectangular shape. A width of the channel may for example be between <NUM> and <NUM>, in some examples between <NUM> and <NUM>. A length of the channel may for example be between <NUM> and <NUM>, in some examples between <NUM> and <NUM>. In another example, the one or more hydrophobic barriers may for example comprise a pair of parallel hydrophobic barriers, each of which may e.g. extend along a straight line to define a pair of opposing walls of the channel.

The one or more hydrophobic barriers may for example comprise wax arranged in the mat of electrically insulating nanofibers, wherein the wax may e.g. surround polymer nanofibers and/or carbon nanofibers in a portion of the mat, e.g. such that nanofibers in the respective portion of the mat are embedded in the wax. In some examples, the wax may fill pores or voids between the nanofibers in the respective portion of the mat at least in part, e.g. such that water can no longer enter or penetrate this portion of the mat.

Forming the one or more hydrophobic barriers may for example comprise arranging a corresponding pattern of wax on the polymer nanofiber mat. The pattern of wax may for example have the same shape as the one or more hydrophobic barriers that are to be formed, i.e. the pattern of wax may comprise a matching structure for each of the one or more hydrophobic barriers. The pattern of wax may for example be printed onto a surface of the polymer nanofiber mat. In other examples, the pattern of wax may be arranged on the polymer nanofiber mat by placing a transfer sheet comprising the pattern of wax on the polymer nanofiber mat. The transfer sheet may for example be a plastic sheet, wherein the pattern of wax may e.g. have been printed onto the plastic sheet prior to placing the transfer sheet on the polymer nanofiber mat. The wax may be melted, e.g. using a hot plate, to transfer the pattern of wax into the polymer nanofiber mat at least in part. A temperature of the hot plate may for example be between <NUM> and <NUM>, in some examples between <NUM> and <NUM>, and the hot plate may e.g. be placed on the transfer sheet for between <NUM> and <NUM> minutes, in some examples between <NUM> and <NUM>.

The pattern of wax on and/or in the transfer sheet may comprise one or more structures having a width between <NUM> and <NUM>, in some examples between <NUM> and <NUM>, preferably between <NUM> and <NUM> in a direction parallel to a surface of the transfer sheet, wherein the width may e.g. be the width prior to placing the transfer sheet onto the polymer nanofiber mat. Preferably, the width is homogenous along the entire length of the respective structures. Additionally or alternative, the width of the one or more structures may be chosen based on the thickness of the polymer nanofiber mat, e.g. such that a ratio between the width w of each of the one or more structures and the thickness d of the polymer mat, w/d, is within a predefined range, for example such that the ratio w/d is between <NUM> and <NUM>, in some examples between <NUM> and <NUM>, in one example between <NUM> and <NUM>, preferably between <NUM> and <NUM>. In a preferred embodiment, all of the structures of the pattern of wax have the same width. In other words, no structure in the pattern of wax may have a physical dimension smaller than a specified width, which may e.g. be between <NUM> and <NUM>.

In other embodiments, the one or more hydrophobic barriers may for example be formed using photolithography, e.g. by soaking the polymer nanofiber mat in a hydrophobic photoresist and structuring the photoresist using light to form the one or more hydrophobic barriers. Additionally or alternatively, the one or more hydrophobic barriers may also be formed by selectively applying a hydrophobic material other than wax to the mat of electrically insulating nanofibers, wherein the hydrophobic material may e.g. be or comprise an oil, glycerol, a hydrophobic glue, a hydrophobic polymer, nail polish or a combination thereof. The hydrophobic material may for example be applied in liquid form, e.g. by screen printing and/or using an automatic dispenser.

Additionally or alternatively, the method may also comprise providing one or more barrier structures other than hydrophobic barriers, wherein the one or more barrier structures may in particular comprise one or more sidewalls of a microfluidic channel, which may e.g. be formed of a water-impermeable material such as plastic, silicone or glass. The one or more sidewalls may in particular be formed in a substrate of a microfluidic chip. Providing the one or more sidewalls of the microfluidic channel may for example comprise placing the polymer nanofiber mat or a part thereof in a microfluidic channel or chamber.

The method may further comprise peeling the polymer nanofiber mat from the substrate or vice-versa, e.g. to obtain a free-standing nanofiber mat or to replace the substrate by a cover sheet. The polymer nanofiber mat may for example be peeled off the substrate using tweezers. Preferably, the polymer nanofiber mat is separated from the substrate after the laser-induced carbonization and/or after forming the one or more hydrophobic barriers. In some embodiments, a cover sheet is placed onto the surface of the polymer nanofiber mat opposite to the substrate prior to separating the polymer nanofiber mat from the substrate, e.g. to provide mechanical stability.

The method may also comprise attaching a cover sheet, in particular a water-impermeable and/or electrically insulating cover sheet, to the polymer nanofiber mat. The cover sheet may for example comprise or consist of glass or plastic, in particular a transparent thermoplastic. In some embodiments, the cover sheet may be the transfer sheet used for forming the one or more hydrophobic barriers. In a preferred embodiment, the method comprises arranging the polymer nanofiber mat between a bottom cover sheet and a top cover sheet, e.g. as detailed below for the sensor according to the invention. In some examples, an adhesive tape, in particular a double-sided adhesive tape, may be placed between the polymer nanofiber mat and one or both of the cover sheets. Additionally or alternatively, one or both cover sheets may also be attached to the polymer nanofiber mat using glue and/or a mechanical fastener such as a clip.

The method may for example be used for forming one or more electrodes for a sensor for performing electrochemical measurements on liquid samples, in particular for the sensor according to the invention described below. The polymer nanofiber mat comprising the one or more carbon nanofiber electrodes may for example be integrated into a microfluidic device, e.g. by placing the polymer nanofiber mat or a part thereof into a microfluidic channel or chamber. A sample fluid may for example flow through the polymer nanofiber mat by capillary transport. Additionally or alternatively, the polymer nanofiber mat comprising the one or more carbon nanofiber electrodes may for example be integrated into a paper-based assay such as a lateral flow test, e.g. by bringing the polymer nanofiber mat in contact with one or more paper strips such as a nitrocellulose strip. A sample fluid may for example flow from a paper strip into the polymer nanofiber mat by capillary transport across the contact interface.

The present invention provides a method of manufacturing a sensor for performing an electrochemical measurement on a liquid sample, wherein the method comprises forming a mat of electrically insulating nanofibers comprising two or more electrically conductive carbon nanofiber electrodes formed therein, the two or more carbon nanofiber electrodes being electrically isolated from each other. The polymer nanofiber mat comprising the two or more carbon nanofiber electrodes may in particular be formed using a method of forming a carbon nanofiber electrode according to any one of the embodiments disclosed herein. The method of manufacturing a sensor further comprises arranging the mat of electrically insulating nanofibers on an electrically insulating bottom cover sheet and providing one or more barrier structures configured to confine the liquid sample to a channel extending through the two or more carbon nanofiber electrodes in the mat of electrically insulating nanofibers.

The polymer nanofiber mat may for example be arranged on the bottom cover sheet as described above for the method of forming a carbon nanofiber electrode. In some embodiments, the bottom cover sheet may be a transfer sheet used for forming one or more hydrophobic wax barriers in the polymer nanofiber mat, e.g. as described above. The method of manufacturing a sensor may further comprise providing an electrically insulating top cover sheet, e.g. to arrange the polymer nanofiber mat with the carbon nanofiber electrodes between the bottom and top cover sheets. The bottom and/or top cover sheets may for example be similar to the bottom and/or top cover sheets of the sensor for performing an electrochemical measurement on a liquid sample according to any one of the embodiments described below.

The one or more barrier structures may for example be similar to the one or more barrier structures of the sensor for performing an electrochemical measurement on a liquid sample according to any one of the embodiments described below and/or may be provided as described above for the method of forming a carbon nanofiber electrode according to the invention. The one or more barrier structures may in particular comprise one or more hydrophobic barriers in the polymer nanofiber mat and/or one or more sidewalls of a microfluidic channel, which may e.g. be formed in a substrate of a microfluidic chip.

The present invention further provides a sensor for performing an electrochemical measurement on a liquid sample. The sensor comprises an electrically insulating bottom cover sheet and a mat of electrically insulating nanofibers arranged on the bottom cover sheet. The electrically insulating nanofibers comprise an organic polymer and two or more electrically conductive carbon nanofiber electrodes are formed in the mat of electrically insulating nanofibers, wherein the two or more carbon nanofiber electrodes are electrically isolated from each other. The sensor further comprises one or more barrier structures configured to confine the liquid sample to a channel extending through the two or more carbon nanofiber electrodes in the mat of electrically insulating nanofibers.

The sensor may in particular be manufactured using a method of forming a carbon nanofiber electrode according to any one of the embodiments described herein and/or using a method of manufacturing a sensor according to any one of the embodiments described herein. Accordingly, the bottom cover sheet, the polymer nanofiber mat, the carbon nanofiber electrodes and the one or more barrier structures may for example be similar to the respective elements described above.

The one or more barrier structures may in particular comprise one or more hydrophobic barriers in the mat of electrically insulating nanofibers, wherein the one or more hydrophobic barriers may define the channel extending through the two or more carbon nanofiber electrodes. The one or more hydrophobic barriers may for example comprise wax surrounding polymer nanofibers and/or carbon nanofibers in the polymer nanofiber mat, e.g. as described above, thereby forming a water-impermeable structure in the polymer nanofiber mat. Additionally or alternatively, the one or more barrier structures may also comprise one or more walls formed of a material that is impermeable for a liquid, in particular water, such as plastic, silicone, and/or glass, wherein the one or more walls may for example define or enclose a microfluidic channel in a substrate. The polymer nanofiber mat comprising the carbon nanofiber electrodes may for example be arranged in a microfluidic channel, e.g. such that the polymer nanofiber mat extends over the entire width and/or a height of the microfluidic channel.

The channel defined by the one or more barrier structures extends within the polymer nanofiber mat and in particular through the carbon nanofiber electrodes therein, for example along a direction parallel to a surface of the polymer nanofiber mat. The one or more barrier structures may in particular be configured to confine the liquid sample in a direction or a plane parallel to a surface of the polymer nanofiber mat. In some embodiments, additional barrier structures such as one or more cover sheets may be arranged above and/or below the polymer nanofiber mat to provide confinement in a direction perpendicular to the surface of the polymer nanofiber mat, e.g. as detailed below. Preferably, the channel extends through some or all of the carbon nanofiber electrodes such that the respective carbon nanofiber electrode covers the entire width and/or the entire height of the channel, e.g. such that the carbon nanofiber electrode extends over the entire cross-section of the channel. In this way, all of the liquid sample flowing along the channel may pass through the carbon nanofiber electrodes.

The sensor may further comprise an electrically insulating top cover sheet, wherein the mat of electrically insulating nanofibers is arranged between the bottom and top cover sheets. One or both of the bottom and top cover sheets preferably comprise a water-impermeable material such as plastic, silicone, and/or glass and may for example be plastic sheets. A thickness of the bottom and/or top cover sheets may for example be between <NUM> and <NUM>, in one example between <NUM> and <NUM>. In some examples, one or both of the bottom and top cover sheets may comprise multiple layers and may for example be a laminate structure.

One or both of the bottom and top cover sheets may comprise one or more openings that are in fluid communication with the channel defined by the one or more barrier structures. The one or more openings may for example comprise an inlet and/or an outlet for the channel. One or both of the bottom and top cover sheets may completely cover the portions of some or all of the two or more carbon nanofiber electrodes arranged within the channel, e.g. such that surfaces of the carbon nanofiber electrode facing the respective cover sheet are not exposed to an environment of the sensor. In other embodiments, one or both of the bottom and top cover sheet may comprise an opening that exposes the portions of some or all of the carbon nanofiber electrodes in the channel, e.g. an opening that extends along the entire channel.

The sensor may also comprise adhesive tape arranged between one or both of the bottom and top cover sheets and the mat of electrically insulating nanofibers. Preferably, the adhesive tape is a double-sided adhesive tape. The adhesive tape may increase the mechanical stability of the sensor and may for example have a thickness between <NUM> and <NUM>, in one example between <NUM> and <NUM>. In some embodiments, the sensor comprises a sandwich structure comprising the bottom cover sheet, an adhesive tape arranged on the bottom cover sheet, the polymer nanofiber mat arranged on the adhesive tape, and a top cover sheet arranged on the polymer nanofiber mat. In one example, the aforementioned layers may be in direct physical contact, i.e. there may be no additional layers arranged between the aforementioned layers.

The organic polymer may be an organic thermoplastic, e.g. as detailed above for the method of forming a carbon nanofiber electrode. The organic polymer may in particular be selected from the group consisting of polyimide, polyacrylonitrile, poly(amic acid), poly(p-xylenetetrahydrothiophenium chloride), polybenzimidazole, poly(vinyl alcohol), and poly(vinylidene fluoride). Preferably, the organic polymer is or comprises polyimide.

The electrically insulating nanofibers and/or the carbon nanofibers in the mat may have properties as described above for the method of forming a carbon nanofiber electrode according to the invention. An average diameter of the electrically insulating nanofibers in the mat may for example be between <NUM> and <NUM>, in some examples between <NUM> and <NUM>. The diameter of the electrically insulating nanofibers may vary between individual nanofibers, wherein the diameters of the electrically insulating nanofibers in the mat may for example be distributed within a range between <NUM> and <NUM>. In some embodiments, the electrically insulating nanofibers and/or the carbon nanofibers are hydrophilic, e.g. as a result of a hydrophilic surface treatment like a plasma treatment as detailed above. In other embodiments, the electrically insulating nanofibers and/or the carbon nanofibers may be hydrophobic, e.g. if the sensor is to be used for measurements on a sample fluid comprising alcohol and/or an amphiphilic substance such as a detergent.

In some embodiments, one or more of the carbon nanofiber electrodes may be functionalized, e.g. as described above. For example, one or more of a catalyst, in particular nanoparticles, e.g. metallic nanoparticles, and/or enzymes, an ion exchanger, in particular a sulfonated tetrafluoroethylene-based fluoropolymer-copolymer, quantum dots, a redox mediator, a conducting polymer, a functional biocompatible polymer, an electrochemically active group, and a functional group configured to bind or immobilize a target substance may be adsorbed, absorbed and/or embedded in one or more of the carbon nanofiber electrodes, e.g. adsorbed, absorbed and/or embedded on or in the carbon nanofibers of the respective electrode.

The sensor may further comprise one or more structures comprising or formed of a porous material, in particular paper, for example one or more paper strips, that are in contact with the polymer nanofiber mat, for example such that the one or more structures are in fluid communication with the channel extending through the two or more carbon nanofiber electrodes, e.g. by capillary transport across the contact interface. This may for example allow for supplying the liquid sample to the channel and in particular for implementing a lateral flow immunoassay. Additionally or alternatively, the sensor may also comprise one or more microfluidic channels that are in fluid communication with the channel extending through the carbon nanofiber electrodes.

In the following, a detailed description of the invention and exemplary embodiments thereof is given with reference to the figures. The figures show schematic illustrations of.

<FIG> shows a flow chart of a method <NUM> of forming a carbon nanofiber electrode according to an exemplary embodiment of the invention. The method <NUM> may for example be used for manufacturing the sensor <NUM> for performing an electrochemical measurement on a liquid sample of <FIG>, which is used as an example for illustrative purposes in the following with steps of the method <NUM> being schematically illustrated in <FIG> (not to scale). This is, however, not intended to be limiting in any way and the method <NUM> may also be used for forming a carbon nanofiber electrode for other purposes and/or for manufacturing a different sensor such as the sensor of <FIG>. Furthermore, the method <NUM> is not limited to the order of execution shown in the flowchart of <FIG>. As far as technically feasible, the method <NUM> may be executed in an arbitrary order and parts thereof may be executed simultaneously at least in part. In particular, step <NUM> or parts thereof may for example be executed prior to, during and/or in between steps <NUM> to <NUM>.

The method <NUM> comprises, in step <NUM>, providing an electrically insulating porous substrate <NUM>. In the example of <FIG>, the substrate <NUM> is a filter paper and is provided on a collector plate <NUM> of a device for electrospinning of nanofibers. The filter paper <NUM> may for example be a grade MN <NUM> filter paper as sold by Macherey-Nagel™ and may for example have a thickness between <NUM> and <NUM>, e.g. <NUM>, wherein the thickness is measured along the Z direction of <FIG>. A diameter of the filter paper <NUM> perpendicular to the Z direction may for example be between <NUM> and <NUM>. The filter paper may for example have an electrical conductivity of about <NUM><NUM> (Ω · m)-<NUM>. Preferably, a plasma treatment is performed on the substrate <NUM> prior to forming a mat <NUM> of electrically insulating nanofibers thereon in step <NUM>, e.g. before placing the substrate <NUM> on the collector plate <NUM>, for example as detailed below for the method of <FIG>.

The method <NUM> further comprises, in step <NUM>, forming a mat <NUM> of electrically insulating nanofibers on the substrate <NUM> by electrospinning, wherein the nanofibers comprise an organic polymer. The electrospinning is for example performed by applying a high voltage between the collector plate <NUM> and a nozzle (not shown) to which a spinning solution comprising the organic polymer is supplied. The voltage may for example be between <NUM> kV and <NUM> kV, in one example between <NUM> kV and <NUM> kV, e.g. <NUM> kV. The nozzle may for example have an opening with a diameter between <NUM> and <NUM>, e.g. <NUM>, and may for example be arranged at a distance of between <NUM> and <NUM>, e.g. <NUM>, above the collector plate <NUM>. The spinning solution may be supplied to the nozzle at a flow rate between <NUM>µl/min and <NUM>µl/min, in some examples between <NUM>µl/min and <NUM>µl/min, e.g. <NUM>µl/min. Ambient parameters such as humidity and temperature may affect properties of the electrospinning nanofibers. The electrospinning may for example be performed at a temperature between <NUM> and <NUM>, preferably between <NUM> and <NUM>, e.g. at <NUM>. The relative humidity may for example be set to between <NUM>% and <NUM>%, in some example between <NUM>% and <NUM>%, preferably between <NUM>% and <NUM>%, e.g. to <NUM>%. To increase the uniformity of the polymer nanofiber mat <NUM>, the substrate <NUM> is rotated, e.g. by <NUM>° after the first half of the electrospinning process.

The spinning solution comprising the organic polymer preferably is an aqueous solution, which may also contain a polar solvent such as dimethylacetamide. The organic polymer may for example be a solvent soluble polyimide such as Matrimid, e.g. Matrimid <NUM>, wherein a concentration of the organic polymer in the spinning solution may for example be between <NUM>/ml and <NUM>/ml, e.g. <NUM>/ml. In addition, the spinning solution may comprise a metal acetylacetonate such as iron (III) acetylacetonate and/or nickel acetylacetonate, e.g. to increase a thermal conductivity of the polymer nanofibers. A concentration of the metal acetylacetonate in the spinning solution may for example be between <NUM>/ml and <NUM>/ml, in one example <NUM>/ml. The present inventors have observed that such a concentration of the metal acetylacetonate in the spinning solution is sufficient for enabling the formation of carbon nanofiber electrodes by laser-induced carbonization without burning the substrate <NUM> and/or the polymer nanofiber mat <NUM>, while higher concentrations, e.g. between <NUM>/ml and <NUM>/ml, did not provide any additional benefits in this regard.

A duration of the electrospinning process may be adapted to control the thickness and/or density of the polymer nanofiber mat <NUM>. A thin polymer nanofiber mat <NUM> may be prone to burning or melting, which may prevent proper carbonization and e.g. lead to a lower electrical conductivity of the carbonized structures. On the other hand, a thick polymer nanofiber mat <NUM> may require higher laser power for carbonization and/or may make performing the method <NUM> more time-consuming. The present inventors have observed that a duration of the electrospinning process between <NUM> and <NUM> and in particular a duration between <NUM> and <NUM> per cm<NUM> of surface area of a collecting area on the substrate <NUM> that is to be coated with polymer nanofibers yields satisfactory polymer nanofiber mats. In one example, the collecting area on the substrate is <NUM> x <NUM> and the electrospinning is performed for between <NUM> and <NUM>, preferably between <NUM> and <NUM>, e.g. <NUM>. The thickness of the resulting polymer nanofiber mat <NUM> may for example be between <NUM> and <NUM>, in some examples between <NUM> and <NUM>.

The method <NUM> further comprises, in step <NUM>, forming one or more electrodes 206A, 206B, 206C comprising electrically conductive carbon nanofibers in the polymer nanofiber mat <NUM> by laser-induced carbonization of polymer nanofibers. For this, a focused laser beam <NUM> is scanned across portions of the polymer nanofiber mat <NUM> that are to be carbonized, which enables the formation of carbon nanofiber electrodes of arbitrary shapes, e.g. as depicted in <FIG>. The carbon nanofiber electrodes 206A-C may for example have physical dimensions in the range of <NUM> to <NUM>, e.g. a width between <NUM> and <NUM> and a length between <NUM> and <NUM>. In some embodiments, a large number of electrodes may be formed in the polymer nanofiber mat <NUM>, e.g. for a plurality of sensors. The laser beam <NUM> may in particular be a pulsed laser beam. A wavelength of the laser beam <NUM> may be in the infrared spectrum and the laser beam <NUM> may for example be generated by a CO<NUM> laser. The laser-induced carbonization process may be optimized by adapting a power, a diameter, a repetition rate, and/or a scanning rate of the laser beam <NUM>, e.g. to achieve the desired degree of carbonization and/or electrical conductivity and/or to prevent burning of the polymer nanofiber mat <NUM> and/or of the substrate <NUM>. A power of the laser beam <NUM> may for example be between <NUM> W and <NUM> W. A scanning rate of the laser beam <NUM> may for example be between <NUM>/s and <NUM>/s. A diameter of the laser beam at its focus may for example be between <NUM> and <NUM>, in one example between <NUM> and <NUM>.

In some embodiments the method <NUM> may further comprise, in step <NUM>, integrating the polymer nanofiber mat <NUM> comprising the carbon nanofiber electrodes 206A-C into a sensor, in particular a sensor for performing an electrochemical measurement on a liquid sample such as the sensor <NUM> of <FIG>. Step <NUM> may for example comprise at least some of the steps of the method of <FIG>.

Manufacturing the sensor <NUM> may in particular comprise forming one or more hydrophobic barriers <NUM> in the polymer nanofiber mat <NUM> to define a channel <NUM> for the liquid sample. In the example of <FIG>, the hydrophobic barriers <NUM> are formed by placing a transfer sheet <NUM> on the polymer nanofiber mat <NUM>, wherein a pattern <NUM> of wax is printed on the transfer sheet <NUM>. The pattern <NUM> of wax is formed to match the pattern of hydrophobic barriers <NUM> that are to be formed in the polymer nanofiber mat <NUM>, e.g. such that the pattern <NUM> of wax comprises a corresponding structure for each of the hydrophobic barriers <NUM>. After placing the transfer sheet <NUM> on the polymer nanofiber mat <NUM>, the wax is melted, e.g. using a hot plate, to transfer the pattern <NUM> of wax into the polymer nanofiber mat <NUM> at least in part. The wax may penetrate the polymer nanofiber mat <NUM> entirely, e.g. such that the wax extends over the entire thickness of polymer nanofiber mat <NUM> from the transfer sheet <NUM> to the substrate <NUM>. The wax may seal off pores and voids between the polymer nanofibers and the carbon nanofibers in the respective portions of the polymer nanofiber mat <NUM> and the carbon nanofiber electrodes 206A-C at least in part, thereby forming hydrophobic barriers that are impenetrable for water, aqueous solutions and/or other liquids. To obtain water-impermeable barriers <NUM> while preventing wax from entering the channel <NUM>, a width or physical dimension of the structures in the pattern <NUM> may for example be chosen to be between <NUM> and <NUM>, preferably between <NUM> and <NUM>, e.g. <NUM>. The width of the structures may for example be between <NUM> and <NUM> times as large as the thickness of the polymer nanofiber mat <NUM>.

In some embodiments, the transfer sheet <NUM>, which may for example be a transparent plastic sheet, may also be used as a top cover sheet <NUM> for the sensor <NUM>, i.e. may remain on the polymer nanofiber mat <NUM> after forming the hydrophobic barriers <NUM>. In the example of <FIG>, the transfer sheet <NUM> comprises a pair of openings 304A, 304B that are arranged on opposite sides of the electrodes 206A-C and provide an inlet and outlet, respectively, for the channel <NUM>.

<FIG> show schematic illustrations (not to scale) of a sensor <NUM> for performing an electrochemical measurement on a liquid sample according to an exemplary embodiment of the invention. The sensor <NUM> is depicted in <FIG> in top view, e.g. along the Z axis of <FIG>, and in <FIG> in a cross-sectional side view, e.g. a cross-section along the line B-B in <FIG>.

The sensor <NUM> comprises a multi-layered substrate <NUM> with an electrically insulating and water-impermeable top cover sheet <NUM>, an electrically insulating and water-impermeable bottom cover sheet <NUM>, a polymer nanofiber mat <NUM> arranged between the top and bottom cover sheets <NUM> and an electrically insulating and water-impermeable adhesive tape <NUM> arranged between the bottom cover sheet <NUM> and the polymer nanofiber mat <NUM>. In the polymer nanofiber mat <NUM>, three electrically conductive carbon nanofiber electrodes 206A-C are arranged, which may e.g. have been formed by a laser-induced carbonization of the polymer nanofiber mat <NUM> as detailed above for the method <NUM>. The carbon nanofiber electrodes 206A-C may for example have an intrinsic electrical conductivity between <NUM> (Ω · m)-<NUM> and <NUM> (Ω · m)-<NUM>. The electrodes 206A-C are electrically isolated from each other, e.g. by non-carbonized portions of the polymer nanofiber mat <NUM>, the top cover sheet <NUM> and the adhesive tape <NUM>. The electrodes 206A-C may for example be used as a working electrode, a counter electrode and a reference electrode in voltammetric measurements on a liquid sample in the channel <NUM>.

Furthermore, a hydrophobic wax barrier <NUM> is formed in the polymer nanofiber mat <NUM>, which encloses a rectangular channel <NUM>. The channel <NUM> extends through each of the three electrodes 206A-C, wherein the electrodes 206A-C cover the entire width and height of the channel <NUM>. In other embodiments, the hydrophobic wax barrier <NUM> may for example be replaced by a hydrophobic barrier formed by photolithography. In yet another example, the hydrophobic wax barrier <NUM> may be replaced by a water-impermeable wall that extends between the adhesive tape <NUM> and the top cover plate <NUM>, wherein the polymer nanofiber mat <NUM> may for example be shaped according to the dimensions of the channel <NUM> to be placed therein, e.g. such that the water-impermeable wall encloses the polymer nanofiber mat <NUM>.

An inlet 304A and an outlet 304B for the channel <NUM> are provided in the top cover sheet <NUM>, wherein a center portion of the top cover sheet <NUM> is in direct contact with upper surfaces of the electrode 206A-C to completely cover at least the portions of the electrodes 206A-C that are arranged within the channel <NUM>. In other embodiments, some or all of the electrodes 206A-C in the channel <NUM> may be exposed at least in part, in some examples completely. The top cover sheet <NUM> may e.g. comprise an opening that extends above the entire channel <NUM> or the sensor <NUM> may not comprise the cover sheet <NUM>.

Each of the electrodes 206A-C comprises a contact pad <NUM>, which is configured to provide an electrical connection to the respective electrode, e.g. through a respective opening or cut-out in the top cover sheet <NUM>. The contact pads <NUM> may for example be formed by portions of the electrodes 206A-C having an enlarged width as illustrated in <FIG>. Additionally or alternatively, the contact pads <NUM> may also comprise other electrically conductive structures such as a metal pad, e.g. a copper pad or foil, and/or a conductive paste, e.g. silver paint. The contact pads <NUM> are arranged outside of the channel <NUM> and may e.g. also be arranged outside of an area of the substrate <NUM> that is covered by the top cover sheet <NUM>.

<FIG> depicts schematic illustrations of various steps of a method of manufacturing a sensor for performing an electrochemical measurement on a liquid sample in accordance with an exemplary embodiment of the invention. Panel a) of <FIG> illustrates the formation of carbon nanofiber electrodes in a polymer nanofiber mat and Panel b) illustrates the integration of the polymer nanofiber mat into a sensor, an exploded view of which is shown in Panel c). Some or all of these steps may for example be executed as part of the method <NUM> of <FIG>, for example in step <NUM>, e.g. to manufacture the sensor <NUM>, which is used as a non-limiting example for illustrative purposes in the following. The method of <FIG> is not limited to the order of execution indicated by the numbering in <FIG>. As far as technically feasible, the method may be executed in an arbitrary order and parts thereof may be executed simultaneously at least in part.

The formation of the carbon nanofiber electrodes comprises (ii) the electrospinning of polymer nanofibers on a filter paper <NUM>, e.g. as in steps <NUM> and <NUM> of method <NUM>, as well as (iii) a selective laser-induced carbonization of the resulting polymer nanofiber mat <NUM>, e.g. as in step <NUM> of method <NUM>.

Prior to forming the polymer nanofiber mat <NUM> on the filter paper <NUM>, a plasma treatment (i) of the filter paper <NUM> may be performed, e.g. to improve adhesion of the polymer nanofiber mat <NUM> on the filter paper <NUM> and/or to prevent curling or bending of the filter paper <NUM> and/or the polymer nanofiber mat <NUM>, which may increase the risk of burning of the filter paper <NUM> and/or the polymer nanofiber mat <NUM> during the laser-induced carbonization. For this, the filter paper <NUM> may be exposed to a plasma, in particular an O<NUM> plasma, preferably a pure (<NUM>%) O<NUM> plasma, wherein the duration of the plasma treatment may for example be between <NUM> and <NUM>, e.g. <NUM>. A power used for generating the plasma may for example be set to between <NUM> Wand <NUM> W, e.g. <NUM> W.

After forming the carbon nanofiber electrodes, a plasma treatment (iv) of the polymer nanofiber mat <NUM> may be performed, e.g. to make the polymer nanofibers and carbon nanofibers hydrophilic for use of the sensor <NUM> with aqueous solutions (NF: polymer nanofibers, LCNF: laser-induced carbon nanofibers). Preferably, an O<NUM> plasma, e.g. a pure (<NUM>%) O<NUM> plasma, is used for this, wherein the duration of the plasma treatment may for example be between <NUM> and <NUM>, e.g. <NUM>. A power used for generating the plasma may for example be set to between <NUM> Wand <NUM> W, e.g. <NUM> W. Such a plasma treatment may for example make the polymer nanofiber mat <NUM> sufficiently hydrophilic to enable complete wetting by aqueous solutions, while at the same time facilitating the formation of water-impermeable barriers such as the hydrophilic wax barriers <NUM>. In some embodiments, the polymer nanofiber mat <NUM> may additionally or alternatively also be treated with an acid such as sulfuric acid. Furthermore, the electrodes 206A-C may be functionalized, e.g. by exposing the electrodes 206A-C to a solution comprising an ion exchanger, for example a solution comprising a cation exchanger such as Nafion, e.g. at a concentration between <NUM>/ml and <NUM>/ml.

Hydrophobic wax barriers <NUM> may be formed in the polymer nanofiber mat <NUM> as shown in Panel b) of <FIG>. The formation of the hydrophobic barriers <NUM> may be similar as described above for step <NUM> of method <NUM> and may comprise (i) printing a corresponding pattern <NUM> of wax onto a plastic transfer sheet <NUM>, (ii) forming inlet and outlet openings 304A, 304B in the transfer sheet <NUM> and (iii) transferring the pattern <NUM> of wax into the polymer nanofiber mat <NUM> to form the hydrophobic barriers <NUM>. For this, the polymer nanofiber mat <NUM> with the transfer sheet <NUM> arranged thereon may for example be placed on a hot plate, e.g. for between <NUM> and <NUM> at a temperature between <NUM> and <NUM>, e.g. <NUM>.

Subsequently, the filter paper <NUM> may be peeled off the polymer nanofiber mat <NUM> (iv), e.g. with tweezers, while the transfer sheet <NUM> remains attached to the polymer nanofiber mat <NUM> to form the top cover sheet <NUM>. The polymer nanofiber mat <NUM> may be turned upside down to place the adhesive tape <NUM> and the bottom cover plate <NUM> thereon (v), thereby closing the sensor <NUM>.

<FIG> contains electron microscope images of a mat of electrically insulating nanofibers comprising a carbon nanofiber electrode formed using a method of forming a carbon nanofiber electrode according to an exemplary embodiment of the invention, e.g. using the method <NUM>. Panel (i) depicts an image of a portion of the polymer nanofiber mat comprising a layer of polyimide nanofibers that were electrospun on a filter paper, wherein a thickness of the polyimide nanofiber mat is <NUM> ± <NUM>. Panel (ii) shows a magnified view of the polyimide nanofibers. The polyimide nanofibers have a diameter of <NUM> ± <NUM> with typical distances between adjacent polyimide nanofibers being in the range of <NUM> to <NUM>. Panel (iii) depicts an image of a portion of the polyimide nanofiber mat that was carbonized using a focused laser beam to form a carbon nanofiber electrode. A thickness of the carbon nanofiber electrode is <NUM> ± <NUM>. A magnified view of the carbon nanofibers is shown in Panel (iv), wherein the scale bar corresponds to <NUM>.

<FIG> shows an image of a sensor for performing an electrochemical measurement on a liquid sample in accordance with an exemplary embodiment of the invention, which has a similar layout as the sensor <NUM> of <FIG>. <FIG> contains electron microscope images of the sensor of <FIG> taken along the lines "A" (Panels A, A1, A2) and "B" (Panel B) of <FIG>, wherein line "A" extends along a channel between an inlet and an outlet of the sensor and line "B" extends along a hydrophobic wax barrier (dark rectangle in <FIG>) surrounding the channel.

The sensor comprises a laminated substrate with plastic sheets as the bottom and top cover sheets, a matrimid nanofiber mat sandwiched between the bottom and top cover sheets and a double-sided adhesive tape arranged between the bottom cover sheet and the matrimid nanofiber mat, cf. Panel A of <FIG>. A total thickness of the substrate is about <NUM>. A channel is formed in the matrimid nanofiber mat by a rectangular hydrophobic wax barrier (dark rectangle in <FIG>). Three carbon nanofiber electrodes formed in the matrimid nanofiber mat extend across the channel with silver paint being provided on contact pads of the electrodes adjacent to the channel to facilitate electrically connecting the sensor to a measurement device.

Panels A1 and A2 of <FIG> show magnified views of the matrimid nanofibers along the channel, which form a water-permeable porous structure through which a liquid sample may flow by capillary transport. Panel B of <FIG> shows an image of the hydrophobic wax barrier defining the channel. Nanofibers in the wax barrier are embedded in wax, thereby forming a water-impermeable wall enclosing the channel.

Claim 1:
A method (<NUM>) of forming a carbon nanofiber electrode (206A-C), the method (<NUM>) comprising:
providing an electrically insulating porous substrate (<NUM>);
forming a mat (<NUM>) of electrically insulating nanofibers on the substrate (<NUM>) by electrospinning, wherein the electrically insulating nanofibers comprise an organic polymer; and
forming an electrode (206A-C) comprising electrically conductive carbon nanofibers in the mat (<NUM>) of electrically insulating nanofibers by laser-induced carbonization of the electrically insulating nanofibers.