Method for producing a device for electrochemical detection of molecules by way of redox cycling, device therefor and the use thereof

Abstract

The invention relates to a method for producing a device for the electrochemical detection of molecules by way of redox cycling, to a device therefor, and to the use thereof. A porous dielectric layer is present between two electrode layers, which is able to receive redox-active molecules and may be biofunctionalized. The individual layers are preferably applied by way of an ink jet printing method.

Claims

1. A device for electrochemical detection of redox-active molecules by way of redox cycling, comprising: a first electrically conductive electrode disposed on a substrate, a passivation layer covering the first electrically conductive electrode at a surface opposite the substrate, the passivation layer having a through-opening; a dielectric layer permeable by said redox-active molecules disposed in the through opening on the first electrically conductive electrode, and a second electrically conductive electrode, having no electrical contact with the first electrode, disposed on the passivation layer and covering the dielectric layer, the second electrically conductive electrode having an access for or being permeable to said redox-active molecules, wherein redox cycling of redox-active molecules permeating the dielectric layer takes place at least at portions of the first electrically conductive electrode and second electrically conductive electrode that are in contact with the dielectric layer, wherein the dielectric layer is a reservoir for the redox-active molecules present in a solution, a pathway for the redox-active molecules to enter the reservoir being comprised by the second electrically conductive electrode through which said redox-active molecules migrate from an exposed surface of the second electrically conductive electrode through the second electrically conductive electrode into the dielectric layer, wherein at least one of the first electrically conductive electrode and second electrically conductive electrode comprises printed electrically conductive particles, and wherein the passivation layer comprises printed electrically insulating particles, and wherein the first electrically conductive electrode and the second electrically conductive electrode of the device are working electrodes in a potentiostat, which are in contact with either one or both of a reference electrode and a counter electrode.

2. A device according to claim 1, wherein the dielectric layer has a surface area between at least 1 m.sup.2 and no more than 1 cm.sup.2.

3. The device according to claim 1 operated by a method comprising: introducing said solution comprising redox-active molecules into the dielectric layer, and applying a voltage to the electrodes so causing alternating reduction and oxidation of the redox-active molecules at the first and second electrically conductive electrodes.

4. The device according to claim 3, wherein said introducing comprises migrating redox-active molecules in solution through pores of the second electrically conductive layer into the dielectric layer.

5. The device according to claim 4, wherein the first electrode has a pore size that is 0 to 50 nm in diameter, the dielectric layer has a pore size that is 10 to 1000 nm in diameter, and the second electrode has a pore size that is 100 to 10000 nm in diameter.

6. The device according to claim 1, wherein the first electrode comprises conductive particles made of gold, platinum, silver, carbon poly(3,4-ethylenedioxythiophene)polystyrene sulfonate or polyaniline; and wherein the second electrode comprises conductive particles made of gold, platinum, silver, carbon, poly(3,4-ethylenedioxythiophene)polystyrene sulfonate or polyaniline.

7. The device according to claim 1, wherein the dielectric layer comprises insulating particles, to which enzymes, antibodies, receptors or other biomolecules can bind.

8. A method for electrochemically detecting redox-active molecules by way of redox cycling comprising: providing a device comprising: a first electrically conductive electrode disposed on a substrate; a passivation layer covering the first electrically conductive electrode at a surface opposite the substrate, the passivation layer having a through-opening; a dielectric layer permeable by said redox-active molecules disposed in the through opening on the first electrically conductive electrode; and a second electrically conductive electrode, having no electrical contact with the first electrode, disposed on the passivation layer and covering the dielectric layer, the second electrically conductive electrode having an access for or being permeable to said redox-active molecules; wherein redox cycling of redox-active molecules permeating the dielectric layer takes place at least at portions of the first electrically conductive electrode and second electrically conductive electrode that are in contact with the dielectric layer; wherein the dielectric layer is a reservoir for the redox-active molecules present in a solution, a pathway for the redox-active molecules to enter the reservoir being comprised by the second electrically conductive electrode through which said redox-active molecules migrate from an exposed surface of the second electrically conductive electrode through the second electrically conductive electrode into the dielectric layer; wherein at least one of the first electrically conductive electrode and second electrically conductive electrode comprises printed electrically conductive particles; wherein the passivation layer comprises printed electrically insulating particles; and wherein the method comprises: introducing said solution comprising redox-active molecules into the dielectric layer, and applying a voltage to the electrodes so causing alternating reduction and oxidation of the redox-active molecules at the first and second electrically conductive electrodes.

9. The method according to claim 8, wherein said introducing comprises migrating redox-active molecules in solution through pores of the second electrically conductive layer into the dielectric layer.

10. The method according to claim 9, wherein the first electrode has a pore size that is 0 to 50 nm in diameter, the dielectric layer has a pore size that is 10 to 1000 nm in diameter, and the second electrode has a pore size that is 100 to 10000 nm in diameter.

11. The device method according to claim 8, wherein the first electrode comprises conductive particles made of gold, platinum, silver, carbon, poly(3,4-ethylenedioxythiophene)polystyrene sulfonate or polyaniline; and wherein the second electrode comprises conductive particles made of gold, platinum, silver, carbon poly(3,4-ethylenedioxythiophene)polystyrene sulfonate or polyaniline.

12. The method according to claim 8, wherein the dielectric layer comprises insulating particles, to which enzymes, antibodies, receptors or other biomolecules can bind.

13. A device for electrochemical detection of redox-active molecules by way of redox cycling, comprising: a first electrically conductive electrode disposed on a substrate; a passivation layer covering the first electrically conductive electrode at a surface opposite the substrate, the passivation layer having a through-opening; a dielectric layer permeable by said redox-active molecules disposed in the through opening on the first electrically conductive electrode; and a second electrically conductive electrode, having no electrical contact with the first electrode, disposed on the passivation layer and covering the dielectric layer, the second electrically conductive electrode having an access for or being permeable to said redox-active molecules; wherein redox cycling of redox-active molecules permeating the dielectric layer takes place at least at portions of the first electrically conductive electrode and second electrically conductive electrode that are in contact with the dielectric layer; wherein the dielectric layer is a reservoir for the redox-active molecules present in a solution, a pathway for the redox-active molecules to enter the reservoir being comprised by the second electrically conductive electrode through which said redox-active molecules migrate from an exposed surface of the second electrically conductive electrode through the second electrically conductive electrode into the dielectric layer; wherein at least one of the first electrically conductive electrode and second electrically conductive electrode comprises printed electrically conductive particles, and wherein the passivation layer comprises printed electrically insulating particles; wherein the second electrically conductive layer is formed with a printable ink comprising electrically conductive particles of a size which are larger than pores in the dielectric layer; and wherein either the first electrically conductive electrode has no pores, or the first electrically conductive electrode has pores that are smaller than a size of dielectric particles in a printable ink used to form the dielectric layer.

14. The device according to claim 13 operated by a method comprising: introducing said solution comprising redox-active molecules into the dielectric layer, and applying a voltage to the electrodes so causing alternating reduction and oxidation of the redox-active molecules at the first and second electrically conductive electrodes.

15. The device according to claim 14, wherein said introducing comprises migrating redox-active molecules in solution through pores of the second electrically conductive layer into the dielectric layer.

16. The device according to claim 15, wherein the first electrode has a pore size that is 0 to 50 nm in diameter, the dielectric layer has a pore size that is 10 to 1000 nm in diameter, and the second electrode has a pore size that is 100 to 10000 nm in diameter.

17. The device according to claim 13, wherein the first electrode comprises conductive particles made of gold, platinum, silver, carbon poly(3,4-ethylenedioxythiophene)polystyrene sulfonate or polyaniline; and wherein the second electrode comprises conductive particles made of gold, platinum, silver, carbon, poly(3,4-ethylenedioxythiophene)polystyrene sulfonate or polyaniline.

18. The device according to claim 13, wherein the dielectric layer comprises insulating particles, to which enzymes, antibodies, receptors or other biomolecules can bind.

Description

IN THE DRAWINGS

(1) FIG. 1 shows a method according to the invention; and

(2) FIG. 2 shows a device according to the invention.

DESCRIPTION OF THE PREFERRED EMBODIMENTS

First Exemplary Embodiment

(3) A sensor comprising sponge-like pores in the second electrode and in the dielectric is produced by way of the above-described method.

(4) Step a): A gold ink is selected as the material for the first electrode 2a. The conducting structures made of the gold ink are printed onto a polyethylene naphthalate (PEN) substrate 1 using an ink jet printer, and are then sintered at 125 C. for 1 hour.

(5) In this way, a first electrode 2a is formed on the substrate 1, which has either no pores or pores having a maximum size of 20 nm.

(6) FIG. 1 shows a right region of the electrode 2a, which defines the active measuring region for the reaction of the redox-active substance (not shown). Furthermore, a left region made of sintered gold ink 2b on the substrate 1 is shown, to which a voltage is applied and which thus represents a conductor track. The region 2a of the first electrode extends to the right out of the image plane and is contacted with a potentiostat to apply voltage. Passivation: An ink made of polyimide is selected. Using this ink, recesses 5*measuring approximately 100100 m2 are defined as electrode regions by way of ink jet printing, as shown in the right part of FIG. 1 (active region). The polyimide ink is disposed thereon so as to passivate the first electrode 2a. In this way, the right active region of the first electrode 2a is passivated around the region 5*. The passivation ink is printed as passivation layers 3a, 3b around the later dielectric 5, so that a region 5*is recessed for the later dielectric 5.

(7) In addition, a portion of the conductor track 2b made of gold is passivated by way of polyimide. In the left inactive region of FIG. 1, the polyimide layer 3c is thus disposed on the conductor track 2b in such a way that the conductor track is partially exposed on the side facing the first electrode 2a, and a step-like arrangement of polyimide 3c and conductor track 2b is produced on the substrate 1.

(8) The passivation layers 3a, 3b and 3c are disposed in a single method step. It goes without saying that the regions of the first electrode 2a and of the conductor track 2b lying in the depth of the image are completely passivated.

(9) Step b) Non-biomodified polystyrene nanoparticle ink comprising nanoparticles 100 nm in size is disposed in the recessed region 5*of the passivation 3a, 3b in the active region of the sensor by way of ink jet printing. This dielectric 5 or this layer 5, due to the porosity thereof, forms a reservoir for the molecule present in solution and to be reacted, or for the analyte/redox mediator. This layer has dimensions of approximately 100 m100 m at a height of 500 nm.

(10) The dielectric 5 is sintered at 115 C. for 5 minutes, so that a homogeneous nanoporous layer 5 is formed as a result of the partial fusion of the particles.

(11) The pore size in the dielectric is approximately 30 nm in diameter.

(12) Step c): Carbon ink comprising carbon nanoparticles of 300 to 400 nm in size is selected as the second, top electrode 4a and disposed on the passivation 3a, 3b and the dielectric 5. The ink is also partially printed onto the passivation layer 3a, 3b and onto the dielectric 5 in the region of the first electrode 2a, and moreover in the inactive region of the sensor, which is shown on the left in FIG. 1, so as to form further contact points for the second electrode in the region 4b via the conductor track 2b. The ink is sintered at 125 C. for 1 hour.

(13) The pore size in the second electrode is approximately 100 nm in diameter.

(14) In addition to the active region for the redox reaction in the right part of FIG. 1, a further particularly advantageous embodiment of the method and of a device thus produced is shown in the left part of FIG. 1. This left region is the so-called inactive region of the sensor. The inactive region comprises the conductor track 2b made of gold, which extends out of the image plane to the left (not shown). The conductor track 2b is contacted with a potentiostat (not shown).

(15) In this way, a voltage, such as above the oxidation potential of the molecule or analyte/redox mediator, can be applied via the conductor track 2b to the active region of the second electrode 4a, which results in oxidation of the molecule/analyte at the electrode. Correspondingly, a voltage is applied to the active region of the first electrode 2a, which is below the reduction potential of the analyte and thus enables the alternating redox cycling process. The detection, however, can also just as well be carried out conversely, so that the reduction potential is applied to the electrode 4a, and the oxidation potential is applied to the electrode 2a.

(16) A molecule or an analyte, such as ferrocenedimethanol, is applied to the electrode in the form of a solution (oxidized or reduced). The bottom, first electrode in the region 2a and the top, second electrode 4a are accordingly brought in contact and set to an oxidizing potential of +600 mV and a reducing potential of 0 mV with respect to an Ag/AgCl reference electrode. The detection of the analyte in various concentrations takes place by measuring the redox cycling current intensity at the oxidizing and/or reducing electrode.

SECOND EXEMPLARY EMBODIMENT: USE OF THE PRINTED REDOX CYCLING SENSOR FOR THE DETECTION OF OVALBUMIN

(17) A second sensor comprising sponge-like pores in the dielectric and the second electrode is produced as follows by way of an above-described method (FIG. 1):

(18) Steps a) and c) and the passivation follow exemplary embodiment 1.

(19) Step a): A gold ink is selected as the material for the first electrode 2a. The conducting structures made of the gold ink are printed onto a polyethylene naphthalate (PEN) substrate 1 using an ink jet printer, and are then sintered at 125 C. for one hour.

(20) In this way, a first electrode 2a is formed on the substrate 1, which has either no pores or pores having a maximum size of 20 nm.

(21) FIG. 1 shows a right region of the electrode 2a, which defines the active measuring region for the reaction of the redox-active substance (not shown). Furthermore, a left region made of sintered gold ink 2b on substrate 1 is shown, which is used for the application of the voltage and thus represents a conductor track. The region 2a of the first electrode extends to the right out of the image plane and is contacted with a potentiostat to apply voltage.

(22) Passivation: An ink made of polyimide is selected. Using this ink, electrode regions approximately 100100 m in size are defined as the recess 5*by way of ink jet printing, as shown in the right part of FIG. 1 (active region). The polyimide ink is provided so as to passivate the first electrode 2a. In this way, the right active region of the first electrode 2a is passivated around the region 5*, this being the subsequent reservoir. The passivation ink is printed as passivation layers 3a, 3b around the later dielectric 5, so that a region 5*is recessed for this dielectric 5.

(23) In addition, a portion of the conductor track 2b made of gold is passivated by way of polyimide. In the left inactive region of FIG. 1, the polyimide layer 3c is thus disposed on the conductor track 2b in such a way that the conductor track is partially exposed on the side facing the first electrode 2a, and a step-like arrangement of polyimide 3c and conductor track 2b is produced on the substrate 1.

(24) The passivation layers 3a, 3b and 3c are disposed in a single method step. It goes without saying that the regions of the first electrode 2a and of the conductor track 2b lying in the depth of the image are completely passivated.

(25) Using this ink, as in exemplary embodiment 1, approximately 100 m100 m electrodes, conductor tracks and contact points for the sensor are again defined.

(26) Step b): 100 nm polystyrene nanoparticle ink, which is to say a polystyrene nanoparticle ink comprising nanoparticles of 100 nm in size, is used. The polystyrene nanoparticles are equipped with anti-ovalbumin antibodies and used as a dielectric 5 or intermediate layer between the two electrodes 2a and 4a. The ink is printed in the region 5*, which was recessed by the passivation layers 3a, 3b so as to define the active region of the electrodes 2a, 4a. The dielectric 5 is heated at 40 C. for 30 minutes, so that the solvents evaporate, but the biological material is not damaged and preserves the function thereof.

(27) The pore size corresponds approximately to that of the first exemplary embodiment.

(28) Step c): Carbon ink comprising carbon nanoparticles of 300 to 400 nm in size is disposed as the top electrode 4a on the passivation layer 3a, 3b and the dielectric 5. The ink is printed onto the passivation layer in the region of the first electrode 2a, and also beyond, so as to form the contact points in the region 4b via the conductor track 2b, and is then sintered, so that the biological material of the dielectric 5 is not damaged.

(29) The pore size corresponds approximately to that of the first exemplary embodiment.

(30) The inactive region thus otherwise corresponds to the inactive region of the first exemplary embodiment, and the contacting of the first electrode 2a and of the generated conductor track 2b beneath the contact region 4b is also identical.

(31) In addition to the active region for the redox reaction in the right part of FIG. 1, a further particularly advantageous embodiment of the method and of a device thus produced is thus shown in the left part of FIG. 1. This left region is the so-called inactive region of the sensor. The inactive region comprises the conductor track 2b made of gold, which extends out of the image plane to the left (not shown). The conductor track 2b is contacted with a potentiostat (not shown).

(32) In this way, a voltage, such as above the oxidation potential of the molecule or analyte/redox mediator, can be applied via the conductor track 2b to the active region of the second electrode 4a above the dielectric 5, which results in oxidation of the molecule/of the analyte at the electrode. Correspondingly, a voltage is applied to the active region of the first electrode 2a which is below the reduction potential of the analyte and thus enables the redox cycling process. The detection can also be carried out conversely, so that the reduction potential is applied to the electrode 4a, and the oxidation potential is applied to the electrode 2a.

(33) A solution comprising ovalbumin and a redox mediator, such as ferrocenedimethanol, is applied to the surface of the second electrode 4a. The bottom electrode 2a and the top electrode 4a are accordingly brought in contact and set to an oxidizing potential of +600 mV and a reducing potential of 0 mV against an Ag/AgCl reference electrode. The detection of the analyte in different concentrations will take place by measuring the redox cycling current intensity.

(34) At higher concentrations of ovalbumin, the redox cycling current intensity will decrease, since the available electrochemical surface also decreases with the concentration.

(35) FIG. 2 shows a schematically illustrated device, which is simplified compared to FIG. 1, in the active region, the cyclical reaction of an analyte at a first electrode 22a or Bot. El. and a second electrode 24a or Top. El. and the arrangement thereof in the potentiostat.

(36) The porous dielectric 25, which serves as the reservoir for the analyte/redox mediator present in solution, is disposed between the two electrodes. Voltages are applied to the porous second electrode 4a and the first electrode 2a, which drive the cyclical redox reaction. The generated current is correspondingly measured against the counter electrode.

(37) According to the invention, a nanoscale redox cycling sensor is thus produced only by way of printing technologies, without additional etching steps or sacrificial layers, and optionally also by way of biomodification without further steps.

(38) The object is achieved by a design comprising electrodes disposed in the Z axis on top of one another, which comprise a nanoscale dielectric between the electrodes, wherein the electrodes and/or the dielectric are completely printed. Advantageously, there are no etching steps in the method. This is achieved in that the three layers (1. first bottom conducting electrode; 2. dielectric layer; 3. second top conducting electrode) have differing porosities. Every further layer has larger particles than the layer disposed beneath, so that the layer 4a lying on top, upon deposition from the liquid phase (such as ink jet printing), cannot flow into the layer 5 lying at the bottom, and cannot flow from layer 5 into layer 2a.

(39) For the exemplary embodiments, an OJ300 ink jet printer from UniJet (Korea) was used.

(40) The Teonex (PEN) substrate was obtained from DuPont-Teijin Films (England).

(41) The Au25 gold ink was obtained from UT Dots (USA).

(42) Polymer inks such as polyimide (PI) PMA-1210P-004 was obtained from Sojitz (Japan). Polystyrene nanoparticle ink was mixed from 200 nm polystyrene beads from Polysciences (USA).

(43) Carbon ink 3800 was obtained from Methode (USA).

(44) Further Exemplary Embodiments:

(45) These relate to the use of the sol gel inks for creating the nanoporous dielectric. In the first exemplary embodiment in step b), for example, the nanoporous dielectric can be provided as follows.

(46) In step b), a non-biomodified sol gel-based ink is prepared. For this purpose, TMOS 1:1:1 (percent by weight) is mixed with deionized water and glycerol in a 100 ml flask and stirred for one hour at room temperature using a magnetic stirrer on a magnetic plate. Afterwards, a 100 mM solution of hydrochloric acid at 500:1 (sol gel:acid, percent by weight) is added for starting the condensation reaction. The sol gel ink is disposed in the recessed region 5*of the passivation 3a, 3b in the active region of the sensor by way of ink jet printing. The dielectric 5 or the layer 5, 25, due to the porosity thereof, forms the reservoir for the molecule present in solution and to be reacted, or for the analyte/redox mediator, after hydrolysis and curing. This layer has dimensions of approximately 100 m100 m at a height of approximately 500 nm.

(47) The dielectric 5, 25 is sintered at room temperature for 60 minutes, so that a homogeneous nanoporous layer 5 forms as a result of the condensation reaction in the printed sol gel layer. The pore size in the dielectric is then approximately 20 to 40 nm in diameter.

(48) It goes without saying that a person skilled in the art can also use other sol gel materials that are subject to an acid-catalyzed and/or base-catalyzed condensation reaction and hydrolysis.

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