WO2011086209A1 - Method for coating electrodes of an electronic device by means of magnetic trapping, resulting electrode, device including same and use of said device - Google Patents

Method for coating electrodes of an electronic device by means of magnetic trapping, resulting electrode, device including same and use of said device Download PDF

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Publication number
WO2011086209A1
WO2011086209A1 PCT/ES2010/070853 ES2010070853W WO2011086209A1 WO 2011086209 A1 WO2011086209 A1 WO 2011086209A1 ES 2010070853 W ES2010070853 W ES 2010070853W WO 2011086209 A1 WO2011086209 A1 WO 2011086209A1
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ntc
electrode
electrodes
modified
ntcs
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PCT/ES2010/070853
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Spanish (es)
French (fr)
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Eva Baldrich Rubio
Fco. Javier MUÑOZ PASCUAL
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Consejo Superior De Investigaciones Científicas (Csic)
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Publication of WO2011086209A1 publication Critical patent/WO2011086209A1/en

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    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B82—NANOTECHNOLOGY
    • B82Y—SPECIFIC USES OR APPLICATIONS OF NANOSTRUCTURES; MEASUREMENT OR ANALYSIS OF NANOSTRUCTURES; MANUFACTURE OR TREATMENT OF NANOSTRUCTURES
    • B82Y15/00—Nanotechnology for interacting, sensing or actuating, e.g. quantum dots as markers in protein assays or molecular motors
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B81—MICROSTRUCTURAL TECHNOLOGY
    • B81C—PROCESSES OR APPARATUS SPECIALLY ADAPTED FOR THE MANUFACTURE OR TREATMENT OF MICROSTRUCTURAL DEVICES OR SYSTEMS
    • B81C1/00—Manufacture or treatment of devices or systems in or on a substrate
    • B81C1/00015—Manufacture or treatment of devices or systems in or on a substrate for manufacturing microsystems
    • B81C1/00206—Processes for functionalising a surface, e.g. provide the surface with specific mechanical, chemical or biological properties
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N27/00—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means
    • G01N27/26—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating electrochemical variables; by using electrolysis or electrophoresis
    • G01N27/28—Electrolytic cell components
    • G01N27/30—Electrodes, e.g. test electrodes; Half-cells
    • G01N27/327—Biochemical electrodes, e.g. electrical or mechanical details for in vitro measurements
    • G01N27/3275—Sensing specific biomolecules, e.g. nucleic acid strands, based on an electrode surface reaction
    • G01N27/3277—Sensing specific biomolecules, e.g. nucleic acid strands, based on an electrode surface reaction being a redox reaction, e.g. detection by cyclic voltammetry
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N27/00—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means
    • G01N27/26—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating electrochemical variables; by using electrolysis or electrophoresis
    • G01N27/28—Electrolytic cell components
    • G01N27/30—Electrodes, e.g. test electrodes; Half-cells
    • G01N27/327—Biochemical electrodes, e.g. electrical or mechanical details for in vitro measurements
    • G01N27/3275—Sensing specific biomolecules, e.g. nucleic acid strands, based on an electrode surface reaction
    • G01N27/3278—Sensing specific biomolecules, e.g. nucleic acid strands, based on an electrode surface reaction involving nanosized elements, e.g. nanogaps or nanoparticles

Definitions

  • the present invention pertains to the field of microelectronic devices and electrode specific coatings of said microelectronic devices.
  • a first object of the present invention is a microelectronic device in which at least one electrode has been modified with a removable coating by magnetic entrapment.
  • a second object of the invention consists in a process for performing removable coatings on electrodes of microelectronic devices by magnetic entrapment.
  • NTC carbon nanotubes
  • NTC modified electrodes The simplest strategy for the production of NTC modified electrodes is to deposit a small volume of a dispersion of NTC (usually prepared in organic solvents) on the electrode surface, followed by evaporation of the solvent.
  • NTC usually prepared in organic solvents
  • a number of alternative strategies have been described to date, based among others on electrodeposition, electrophoresis, stamping, spin-coating, entrapment, chemical / covalent conjugation and self-assembling of alternate layers (Layer-by-layer formation) of the NTC.
  • the NTCs are incorporated into the electrode surface in combination with biopolymers, mineral oils, conductive polymers and / or nanoparticles, shaping a variety of composite materials that significantly improve the electrochemical performance of the electrode.
  • NTC carbon nanotubes
  • NTC dispersed in organic solvents has in some cases detrimental effects on the integrity of the electrodes, for example the dissolution of the pastes / inks with which the screen-printed electrodes (SPE) are produced and the reduction of the average life time of the same.
  • the process object of the invention describes an extremely fast and simple method for the reversible production of modified electronic or microelectronic devices with carbon nanotube (NTC) coatings by magnetic entrapment in aqueous medium.
  • NTC carbon nanotube
  • NTC Carbon nanotubes
  • NTC carbon nanotubes
  • NTCs that have undergone a treatment (physical / chemical) that makes them hydrophilic / water soluble (for example, but not exclusively, generation of surface carboxyl groups by acid treatment), or NTCs that have been chemically modified to 5 incorporate hydrophilic / water-soluble groups (for example, but not exclusively, chemical incorporation of ethylene glycol groups).
  • the NTCs are adsorbed nonspecifically on the surface of magnetic microparticles (PM).
  • PM magnetic microparticles
  • PM can be PM of 1 to several microns in diameter, regardless of their origin, composition or manufacturing process, as long as they are soluble in aqueous medium.
  • PM may be protein coated PM, but PM with other surfaces / coatings could also be used (for example, but not exclusively, positively or negatively charged PM, polymer coated PM, metal coated PM, or PM exhibiting surface reactive chemical groups).
  • the process may include, in addition to or as an alternative to the nonspecific adsorption of the NTCs, their chemical conjugation, or the stabilization of the PM / NTC complexes by any other means.
  • PM and / or NTC modified with reactive groups can be used (for example, but not exclusively, PM or NTC that exhibit succinimido groups, biotin molecules, etc., on the surface, so that they react with NH 2 - groups, molecules of streptavidite, etc., present or incorporated on the surface of NTC or PM), or chemically conjugate PM and NTC in the presence of suitable reagents.
  • the device to be modified must incorporate, for example below or in its vicinity, a magnet or a magnetic field generator of any type. This should ensure that the PM / NTC complexes are deposited directly 0 on the sensor surface, working electrode, etc.
  • the invention encompasses devices of any size, geometry, shape or material. It can be flat geometry devices, such as, but not exclusively, screen-printed electrodes and / or microelectrodes manufactured using silicon technology.
  • the process object of the invention is capable of being applicable to different devices, such as electrodes of carbon paste, graphite, NTC or similar, which may include in its composition other compounds (polymers, adhesives, conductive materials, etc) or nanomaterials (nanoparticles / nanowires / ect. of any material), and that can integrate a magnet or the like.
  • the magnet is reversibly incorporated. That is, after the deposition of the PM / NTC and / or the taking of measures can be withdrawn.
  • the procedure consists of mixing the selected volumes of both components.
  • the relative amounts and / or proportions of both components may be varied and will depend on the application.
  • the mixture is deposited (immediately or after incubation) on at least one of the electrodes of the device (for example, but not exclusively, a working electrode) that wants to be modified with NTC and that is being subjected to the effect of a magnet or a magnetic field generator located on the underside of the electrode.
  • the volume or amount of PM / NTC deposited may be varied and will depend on the size, geometry and characteristics of the device to be modified, as well as the intended application. Thanks to the effect of the magnetic field, PM / NTCs are deposited on the surface of the device and in contact with it. This sediment can be subjected to a subsequent wash with the desired aqueous / saline solutions.
  • the surface modified by the described procedure must not be dried.
  • the modified device can be used for the taking of any type of electrochemical measurements including, but not exclusively, amperometric, voltammetric, impedimetric and / or conductimetric.
  • Figure 1 It shows a graph of the current generated in unmodified electrodes (bare SPE) or modified with 5 ⁇ of PM and increasing amounts of NTC (0-7.5 ⁇ NTC, 1 mg / ml). Cyclic voltammetry in 250 ⁇ of ferrocyanide (dark bars) or in PBS (background current, light bars).
  • Figure 2 Shows a graph of the current generated in the unmodified electrodes (bare SPE) or modified with 5 ⁇ of NTC and increasing amounts of PM (1 -10 ⁇ ). Cyclic voltammetry in 250 ⁇ of ferrocyanide (dark bars) or in PBS (background current, light bars).
  • Figure 3 It shows cyclic voltamograms obtained for increasing concentrations of dopamine (DA), dissolved in PBS, on an unmodified electrode.
  • Figure 4 It shows cyclic voltamograms obtained for increasing concentrations of DA, dissolved in PBS, on an electrode modified with 10 ⁇ of PM and 5 ⁇ of NTC deposited by magnetic entrapment.
  • Figure 5 Shows a graph with the peak current values obtained for increasing concentrations of DA in modified electrodes and without Modify.
  • Figure 6. Shows a graph of a difference pulse voltammetry (DPV) obtained in dopamine (DA) and uric acid (AU) measured on an unmodified electrode.
  • Figure 7. Shows a graph of a difference pulse voltammetry (DPV) obtained in dopamine (DA) and uric acid (AU) measured on a modified electrode with PM / NTC.
  • Single wall NTC (SWNTC) modified with carboxyl groups (-COOH) on their surface were used.
  • the NTCs were resuspended in Phosphate Buffered Saline (PBS) at a final concentration of 1 mg / ml and dispersed by sonication for at least 30 minutes at room temperature. The suspension was then stirred using a vortex for 2 minutes.
  • PBS Phosphate Buffered Saline
  • These NTCs are soluble in aqueous medium up to 1 mg / ml; they have an average individual diameter of 1.4 nm ⁇ 0.1 nm and form beams of 4-5 nm 0.5-1.5 microns.
  • the NTCs were sonicated for 15 minutes, followed by vortexing for another 1 minute.
  • the PM were 2.8 microns in diameter and were coated with protein.
  • the MPs were received at an approximate concentration of E 3-3.5 x 10 8 particles per milliliter (equivalent to 3-3.5 x 10 5 MP, approximately 5 mg, per microliter).
  • the MPs were agitated using a vortex for one minute, to completely resuspend them before use, and the necessary volume was transferred to an Eppendorf tube.
  • the PM were concentrated with the help of a magnet or a first magnetic field generator, the supernatant was removed, PM were washed 3 times with PBS (10mM Phosphate Buffer Saline, pH 7.4) and resuspended in PBS at the chosen concentration.
  • Electrodes consisting of flat screen-printed electrodes composed of a working electrode (1.6 mm in diameter) and a gold auxiliary electrode, and an Ag / AgCI reference electrode, all of them printed on a ceramic substrate, were used. Prior to use, the electrodes were electrochemically activated by cyclic voltammetry between 0 and 1.4 V in 1 MH 2 S0 4 until a stable signal was obtained. Subsequent characterization was carried out by cyclic voltammetry in 0.1 M K3 [Fe (CN) 6].
  • a 1 mm diameter cylindrical magnet was fixed under the working electrode as a second magnetic field generator.
  • the second magnetic field generator is removed from the back of the working electrode to release the previously produced coating on the working electrode; leaving the device in its original state.
  • EXAMPLE 1 Efficiency of electrodes modified by magnetic entrapment of increasing amounts of NTC.
  • the entrapment of MP without NTC generates partial passivation of the electrode, increase in the background current in PBS and decrease in the current recorded in ferrocyanide.
  • entrapment of increasing amounts of NTC results in an additional increase in the background current in PBS, whose value is directly related to the amount of NTCs captured, and induces the appearance of a pre-peak between -0.1 and 0 V.
  • entrapment of increasing amounts of NTC correlates with a linear increase in the signal recorded in ferricyanide for amounts between 1 and 5 ⁇ g of NTC.
  • the reagents, electrodes and experimental procedure are similar to those described in example 1.
  • EXAMPLE 3 Detection of dopamine (DA) using electrodes modified by magnetic entrapment of PM / NTC.
  • the third example includes the detection of an electroactive molecule of clinical interest, dopamine (DA), using electrodes modified with PM / NTC by magnetic entrapment.
  • DA is a neurotransmitter released by the hypothalamus and the malfunction of its activity has been linked to several neurodegenerative disorders, including anorexia and bulimia, Parkinson's and Alzheimer's diseases, and schizophrenia.
  • the reagents, electrodes and experimental procedure are similar to those described in example 1. In this case the surface was modified with 3 x 10 6 PM resuspended in 5 ⁇ g of NTC.
  • DA is not very reversible, because the oxidized product adsorbs on the electrode surface.
  • the recorded bottom currents are higher, in line with the increase in the electrode surface due to the deposition of the NTC.
  • the separation between peaks is 0.10 V for the modified electrode, compared to 0.15 V for the unmodified electrode.
  • the oxidation peak current increases with the concentration of DA.
  • the reversibility of the reaction improves on the electrodes modified with NTC, and so does the detectability of DA.
  • the recorded signals are, for all tested DA concentrations, 2.6 to 5 times higher in the modified electrodes than the signals generated in the unmodified electrodes.
  • Figure 5 shows the average data, calculated from the results of three independent electrodes.
  • the detection limit calculated as the average of at least five targets plus three times the standard deviation thereof, improves from 1.69 ⁇ 69 in the unmodified electrodes to 0.71 ⁇ when electrodes with PM / NTC are used.
  • the sensitivity of the test measured as the slope of the linear part of the graph, also improves on surface modified with PIW NTC (30 vs 9 nAmp / ⁇ DA).
  • EXAMPLE 4 Simultaneous detection of dopamine and uric acid using electrodes modified by magnetic entrapment of PM / NTC.
  • AD uric acid
  • AU uric acid
  • the AU oxidizes in most electrodes to potentials so close to those of the DA that the peaks generated by both compounds are hardly distinguishable.
  • NTC uric acid
  • the incorporation of NTC to the electrode usually improves the resolution of the peaks generated by the DA and the AU.
  • DUV Differential Pulse Voltammetry

Abstract

The invention relates to a method for producing coatings on one of the electrodes, known as the working electrode, of electronic devices used to take amplified electrochemical measurements. According to the invention, once the measurement has been taken, the coating can be removed, returning the electronic device to its pre-coating state, thereby allowing the re-use thereof for other measurements, using the same or a different coating.

Description

PROCEDIMIENTO DE RECUBRIMIENTO DE ELECTRODOS DE UN DISPOSITIVO ELECTRÓNICO POR ATRAPAMIENTO MAGNÉTICO, ELECTRODO ASI OBTENIDO, DISPOSITIVO QUE INCORPORA DICHO  PROCEDURE FOR COATING ELECTRODES OF AN ELECTRONIC DEVICE BY MAGNETIC ATTACKING, ELECTRODE SO OBTAINED, DEVICE THAT INCLUDES SUCH
ELECTRODO Y USO DE DICHO DISPOSITIVO  ELECTRODE AND USE OF SUCH DEVICE
D E S C R I P C I Ó N D E S C R I P C I Ó N
OBJETO DE LA INVENCIÓN OBJECT OF THE INVENTION
La presente invención pertenece al campo de los dispositivos microelectrónicos y los recubrimientos específicos para electrodos de dichos dispositivos microelectrónicos.  The present invention pertains to the field of microelectronic devices and electrode specific coatings of said microelectronic devices.
Un primer objeto de la presente invención es un dispositivo microelectrónico en que al menos un electrodo ha sido modificado con un recubrimiento desprendible mediante atrapamiento magnético.  A first object of the present invention is a microelectronic device in which at least one electrode has been modified with a removable coating by magnetic entrapment.
Un segundo objeto de la invención consiste en un procedimiento para poder realizar recubrimientos desprendibles sobre electrodos de dispositivos microelectrónicos mediante atrapamiento magnético.  A second object of the invention consists in a process for performing removable coatings on electrodes of microelectronic devices by magnetic entrapment.
ANTECEDENTES DE LA INVENCIÓN BACKGROUND OF THE INVENTION
Desde su descubrimiento, los nanotubos de carbono (NTC) se han convertido en uno de los nanomateriales más prometedores y tanto el procedimiento de producción como su purificación, modificación y/o funcionalización han sido extensamente sometidos a protección industrial. Aparte de otras muchas aplicaciones potenciales, los NTC han sido explotados en un importante número de aplicaciones electroanalíticas y sensoras. Esto es principalmente debido al hecho de que la incorporación de NTC a los diferentes tipos de electrodos permite tomar ventaja de la alta resistencia mecánica de este nanocomponente, que también muestra gran estabilidad química y conductividad electrónica. Los electrodos modificados con NTC presentan además superficies activas de mayor rugosidad y superficie, niveles más bajos de adsorción inespecífica de biocomponentes, niveles importantes de actividad electrocatalítica hacia una gran variedad de moléculas, y mayor eficiencia de transferencia de electrones que electrodos similares no modificados. Since its discovery, carbon nanotubes (NTC) have become one of the most promising nanomaterials and both the production process and its purification, modification and / or functionalization have been extensively subjected to industrial protection. Apart from many other potential applications, NTCs have been exploited in a significant number of electroanalytical and sensor applications. This is mainly due to the fact that the incorporation of NTC to the different types of electrodes allows to take advantage of the high mechanical resistance of this nanocomponent, which also shows great chemical stability and electronic conductivity. NTC modified electrodes also have active surfaces of greater roughness and surface, lower levels of nonspecific adsorption of biocomponents, significant levels of electrocatalytic activity towards a great variety of molecules, and greater efficiency of electron transfer than similar unmodified electrodes.
La estrategia más simple para la producción de electrodos modificados con NTC consiste en depositar un pequeño volumen de una dispersión de NTC (por lo general preparada en disolventes orgánicos) sobre la superficie del electrodo, seguido de la evaporación del disolvente. Sin embargo, un cierto número de estrategias alternativas han sido descritas hasta la fecha, basadas entre otras en la electrodeposición, electroforesis, estampado, spin-coating, atrapamiento, conjugación química/covalente y autoensamblado de capas alternas (Layer-by- layer formation) de los NTC. En parte de los trabajos existentes, los NTC se incorporan a la superficie del electrodo en combinación con biopolímeros, aceites minerales, polímeros conductores y/o nanopartículas, dando forma a una variedad de materiales compuestos que mejoran de forma importante el rendimiento electroquímico del electrodo. The simplest strategy for the production of NTC modified electrodes is to deposit a small volume of a dispersion of NTC (usually prepared in organic solvents) on the electrode surface, followed by evaporation of the solvent. However, a number of alternative strategies have been described to date, based among others on electrodeposition, electrophoresis, stamping, spin-coating, entrapment, chemical / covalent conjugation and self-assembling of alternate layers (Layer-by-layer formation) of the NTC. In part of the existing works, the NTCs are incorporated into the electrode surface in combination with biopolymers, mineral oils, conductive polymers and / or nanoparticles, shaping a variety of composite materials that significantly improve the electrochemical performance of the electrode.
Sin embargo, estos procedimientos generalmente implican estrategias de producción de al menos horas y rara vez son compatibles con la regeneración fácil del electrodo mediante eliminación de los NTC. Un solo informe (patente y artículo 2004-2005) describe el uso combinado de NTC y nanopartículas magnéticas. En él se describe la adsorción inespecíficas de nanopartículas magnéticas (5-100 nm) sobre la superficie de los NTC, suspensión en una matriz (polimérica, cerámica, metálica, un gel), alineamiento mediante un campo magnético, y solidificación del compuesto por secado/eliminación de la matriz. El material resultante es entonces cortado en láminas, sin especificarse ninguna aplicación potencial. However, these procedures generally involve production strategies of at least hours and are rarely compatible with the easy regeneration of the electrode by removing the NTCs. A single report (patent and article 2004-2005) describes the combined use of NTC and magnetic nanoparticles. It describes the nonspecific adsorption of magnetic nanoparticles (5-100 nm) on the surface of the NTC, suspension in a matrix (polymeric, ceramic, metallic, a gel), alignment by a magnetic field, and solidification of the compound by drying / removal of the matrix. The resulting material is then cut into sheets, without specifying any potential application.
La incorporación de nanotubos de carbono (NTC) a la superficie de electrodos contribuye a aumentar su rugosidad y superficie, reduce el nivel de adsorción inespecífica de biocomponentes, proporciona actividad electrocatalítica frente a una variedad de moléculas, y mejora la transferencia de electrones. Hasta la fecha, este tipo de modificación se basa en la deposición irreversible de los NTC sobre la superficie mediante estrategias poco compatibles con la reutilización del electrodo, a menudo utilizando disolventes orgánicos potencialmente dañinos para el operador y el medio ambiente. Además, como los NTC son un material altamente poroso, los electrodos modificados con NTC pueden sufrir niveles de adsorción inespecífica y/o electrodeposición de biomoléculas demasiado altos para garantizar ausencia de contaminación cruzada entre muestras en estudio y la especificidad y reproducibilidad del protocolo de medida. La deposición de NTC dispersos en disolventes orgánicos tiene en algunos casos efectos perjudiciales en la integridad de los electrodos, por ejemplo la disolución de las pastas/tintas con las que se producen los electrodos serigrafiados (SPE) y la reducción del tiempo de vida media de los mismos. DESCRIPCIÓN DE LA INVENCIÓN The incorporation of carbon nanotubes (NTC) to the electrode surface contributes to increase its roughness and surface area, reduces the level of nonspecific adsorption of biocomponents, provides electrocatalytic activity against a variety of molecules, and improves electron transfer. To date, this type of modification is based on the Irreversible deposition of NTCs on the surface through strategies that are poorly compatible with electrode reuse, often using organic solvents potentially harmful to the operator and the environment. In addition, since NTCs are a highly porous material, NTC modified electrodes can suffer from unspecific adsorption levels and / or electrodeposition of biomolecules that are too high to guarantee the absence of cross contamination between samples under study and the specificity and reproducibility of the measurement protocol. The deposition of NTC dispersed in organic solvents has in some cases detrimental effects on the integrity of the electrodes, for example the dissolution of the pastes / inks with which the screen-printed electrodes (SPE) are produced and the reduction of the average life time of the same. DESCRIPTION OF THE INVENTION
El procedimiento objeto de la invención describe un método extremadamente rápido y sencillo para la producción reversible de dispositivos electrónicos o microelectrónicos modificados con recubrimientos de nanotubos de carbono (NTC) mediante atrapamiento magnético en medio acuoso. The process object of the invention describes an extremely fast and simple method for the reversible production of modified electronic or microelectronic devices with carbon nanotube (NTC) coatings by magnetic entrapment in aqueous medium.
Dicho procedimiento explota la fuerte tendencia que muestran los NTC a adsorberse inespecíficamente sobre la superficie de partículas magnéticas (PM), por ejemplo PM que han sido previamente recubiertas con proteína. Los ensamblados de PM/NTC son subsiguientemente capturados sobre la superficie de un dispositivo electrónico o microelectrónico mediante captura magnética, un procedimiento que resulta extremadamente rápido y sencillo. Los dispositivos modificados de esta forma presentan comportamiento electrocatalítico frente a una variedad de analitos, mejores límites de detección y señales significativamente más altas que los electrodos sin modificar. Tras la toma de medidas, y en ausencia de atracción magnética, los PM/NTC pueden ser eliminados fácilmente mediante lavado y la superficie del dispositivo puede ser reutilizada indefinidamente. This procedure exploits the strong tendency shown by NTCs to adsorb unspecifically on the surface of magnetic particles (PM), for example PM that have been previously coated with protein. PM / NTC assemblies are subsequently captured on the surface of an electronic or microelectronic device by magnetic capture, a procedure that is extremely fast and simple. Devices modified in this way exhibit electrocatalytic behavior against a variety of analytes, better detection limits and significantly higher signals than unmodified electrodes. After taking measurements, and in the absence of magnetic attraction, PM / NTC can be easily removed by washing and the surface of the device can be reused indefinitely.
Los nanotubos de carbono (NTC) han sido explotados para un número importante de aplicaciones electroanalíticas y sensoras. Su incorporación a la superficie de electrodos contribuye a aumentar su rugosidad y superficie, reduce el nivel de adsorción inespecífica de biocomponentes, proporciona actividad electrocatalítica frente a una variedad de moléculas, y mejora la transferencia de electrones. Hasta la fecha, este tipo de modificación se basa en la deposición irreversible de los NTC sobre la superficie mediante estrategias que son poco compatibles con la reutilización del electrodo. No obstante, los NTC son un material altamente poroso y los electrodos modificados con NTC son susceptibles de promover niveles importantes de adsorción inespecífica y/o electrodeposición de biomoléculas y/o biomoléculas, lo que podría inducir contaminación cruzada entre muestras en estudio y afectar la especificidad y reproducibilidad del protocolo de medida. Este inconveniente ha sido a menudo eludido mediante la combinación de los NTC con polímeros cargados eléctricamente capaces de repeler las moléculas de carga negativa / positiva. Carbon nanotubes (NTC) have been exploited for a significant number of electroanalytical and sensor applications. Its incorporation to the electrode surface helps to increase its roughness and surface area, reduces the level of nonspecific adsorption of biocomponents, provides electrocatalytic activity against a variety of molecules, and improves electron transfer. To date, this type of modification is based on the irreversible deposition of NTCs on the surface through strategies that are poorly compatible with electrode reuse. However, NTCs are a highly porous material and NTC modified electrodes are likely to promote significant levels of nonspecific adsorption and / or electrodeposition of biomolecules and / or biomolecules, which could induce cross contamination between samples under study and affect specificity. and reproducibility of the measurement protocol. This drawback has often been eluded by combining NTCs with electrically charged polymers capable of repelling negative / positive charge molecules.
Por otra parte usando NTC carboxilados (con grupos carboxilo en superficie) dispersos en medio acuoso, se observa una fuerte tendencia de los mismos a adsorberse inespecíficamente sobre a superficies recubiertas de proteína. En esta invención se hace uso de esta capacidad para producir PM recubiertas de NTC y la subsiguiente captura magnética de estos complejos de PM/NTC para la producción de electrodos modificados con NTC. El protocolo de producción resulta extremadamente rápido, fácil de llevar a cabo, y no requiere la utilización de disolventes orgánicos evitando efectos perjudiciales en la integridad de los electrodos. Esto minimiza la exposición a los disolventes y a su vez resulta beneficioso para el medio ambiente a nivel de gestión de residuos. El procedimiento descrito en esta invención se basa en la utilización de nanotubos de carbono (NTC), independientemente de su origen, método de producción, composición o características, siempre y cuando sean solubles en medio acuoso. Puede tratarse de NTC que hayan sido sometidos a un tratamiento (físico/químico) que los haga hidrofílicos/hidrosolubles (por ejemplo, pero no exclusivamente, generación de grupos carboxilo en superficie mediante tratamiento ácido), o de NTC que hayan sido modificados químicamente para 5 incorporar grupos hidrofílicos/hidrosolubles (por ejemplo, pero no exclusivamente, incorporación química de grupos etilenglicol). On the other hand, using carboxylated NTCs (with surface carboxyl groups) dispersed in aqueous medium, a strong tendency is observed to adsorb nonspecifically on protein-coated surfaces. In this invention, this capacity is used to produce NTC coated PM and the subsequent magnetic capture of these PM / NTC complexes for the production of NTC modified electrodes. The production protocol is extremely fast, easy to carry out, and does not require the use of organic solvents avoiding harmful effects on the integrity of the electrodes. This minimizes exposure to solvents and in turn is beneficial to the environment at the level of waste management. The procedure described in this invention is based on the use of carbon nanotubes (NTC), regardless of their origin, production method, composition or characteristics, provided they are soluble in aqueous medium. These can be NTCs that have undergone a treatment (physical / chemical) that makes them hydrophilic / water soluble (for example, but not exclusively, generation of surface carboxyl groups by acid treatment), or NTCs that have been chemically modified to 5 incorporate hydrophilic / water-soluble groups (for example, but not exclusively, chemical incorporation of ethylene glycol groups).
En este procedimiento, los NTC son adsorbidos inespecíficamente sobre la superficie de micropartículas magnéticas (PM). Puede tratarse de PM de 1 a o varias mieras de diámetro, independientemente de su procedencia, composición o proceso de fabricación, siempre y cuando sean solubles en medio acuoso. Dichas PM pueden ser PM recubiertas de proteína, pero PM con otras superficies/recubrimientos podrían ser también utilizadas (por ejemplo, pero no exclusivamente, PM cargadas positiva o negativamente, PM recubiertas de5 polímero, PM recubiertas de una capa metálica, o PM que exhiban en superficie grupos químicos reactivos). In this procedure, the NTCs are adsorbed nonspecifically on the surface of magnetic microparticles (PM). It can be PM of 1 to several microns in diameter, regardless of their origin, composition or manufacturing process, as long as they are soluble in aqueous medium. Such PM may be protein coated PM, but PM with other surfaces / coatings could also be used (for example, but not exclusively, positively or negatively charged PM, polymer coated PM, metal coated PM, or PM exhibiting surface reactive chemical groups).
El procedimiento puede incluir, además de o como alternativa a la adsorción inespecífica de los NTC, su conjugación química, o la estabilización de o los complejos de PM/NTC por cualquier otro medio. Para ello se puede utilizar PM y/o NTC modificados con grupos reactivos (por ejemplo, pero no exclusivamente, PM o NTC que exhiban en superficie grupos succinimido, moléculas de biotina, etc., para que éstos reaccionen con grupos NH2-, moléculas de estreptavidita, etc., presentes o incorporados en la superficie de NTC o PM), o conjugar 5 químicamente PM y NTC en presencia de reactivos adecuados. The process may include, in addition to or as an alternative to the nonspecific adsorption of the NTCs, their chemical conjugation, or the stabilization of the PM / NTC complexes by any other means. For this, PM and / or NTC modified with reactive groups can be used (for example, but not exclusively, PM or NTC that exhibit succinimido groups, biotin molecules, etc., on the surface, so that they react with NH 2 - groups, molecules of streptavidite, etc., present or incorporated on the surface of NTC or PM), or chemically conjugate PM and NTC in the presence of suitable reagents.
El dispositivo a modificar debe incorporar, por ejemplo debajo o en su proximidad, un imán o un generador de campo magnético de cualquier tipo. Este debe garantizar que los complejos de PM/NTC son depositados directamente 0 sobre la superficie sensora, electrodo de trabajo, etc. La invención engloba dispositivos de cualquier tamaño, geometría, forma o material. Puede tratarse de dispositivos de geometría plana, como por ejemplo, pero no exclusivamente, electrodos serigrafiados y/o microelectrodos fabricados mediante tecnología de silicio. Sin embargo el procedimiento objeto de la invención es susceptible de ser aplicable a dispositivos diferentes, como los electrodos de pasta de carbono, grafito, NTC o similar, que pueden incluir en su composición otros compuestos (polímeros, materiales adhesivos, materiales conductores, etc) o nanomateriales (nanopartículas/nanohilos/ect. de cualquier material), y que puedan integrar un imán o similar. The device to be modified must incorporate, for example below or in its vicinity, a magnet or a magnetic field generator of any type. This should ensure that the PM / NTC complexes are deposited directly 0 on the sensor surface, working electrode, etc. The invention encompasses devices of any size, geometry, shape or material. It can be flat geometry devices, such as, but not exclusively, screen-printed electrodes and / or microelectrodes manufactured using silicon technology. However, the process object of the invention is capable of being applicable to different devices, such as electrodes of carbon paste, graphite, NTC or similar, which may include in its composition other compounds (polymers, adhesives, conductive materials, etc) or nanomaterials (nanoparticles / nanowires / ect. of any material), and that can integrate a magnet or the like.
En el supuesto de querer reutilizar los dispositivos tras la toma de medidas mediante eliminación de los complejos de PM/NTC, el imán se incorpora de forma reversible. Es decir, tras la deposición de los PM/NTC y/o la toma de medidas puede ser retirado. In the case of wanting to reuse the devices after taking measurements by eliminating the PM / NTC complexes, the magnet is reversibly incorporated. That is, after the deposition of the PM / NTC and / or the taking of measures can be withdrawn.
Para la producción de los complejos de PM/NTC y la modificación de los dispositivos, el procedimiento consiste en la mezcla de los volúmenes seleccionados de ambos componentes. Las cantidades y/o proporciones relativas de ambos componentes pueden ser variadas y dependerán de la aplicación. La mezcla se deposita (inmediatamente o después de incubación) sobre al menos uno de los electrodos del dispositivo (por ejemplo, pero no exclusivamente, un electrodo de trabajo) que quiere ser modificado con NTC y que está siendo sometido al efecto de un imán o un generador de campo magnético ubicado en el envés del electrodo. El volumen o cantidad de PM/NTC depositado podrá ser variado y dependerá del tamaño, geometría y características del dispositivo a modificar, así como de la aplicación prevista. Gracias al efecto del campo magnético, los PM/NTC son depositados sobre la superficie del dispositivo y en contacto con la misma. Este sedimento puede someterse a un lavado posterior con las soluciones acuosas/salinas deseadas. For the production of the PM / NTC complexes and the modification of the devices, the procedure consists of mixing the selected volumes of both components. The relative amounts and / or proportions of both components may be varied and will depend on the application. The mixture is deposited (immediately or after incubation) on at least one of the electrodes of the device (for example, but not exclusively, a working electrode) that wants to be modified with NTC and that is being subjected to the effect of a magnet or a magnetic field generator located on the underside of the electrode. The volume or amount of PM / NTC deposited may be varied and will depend on the size, geometry and characteristics of the device to be modified, as well as the intended application. Thanks to the effect of the magnetic field, PM / NTCs are deposited on the surface of the device and in contact with it. This sediment can be subjected to a subsequent wash with the desired aqueous / saline solutions.
Para garantizar la reutilización de los dispositivos mediante eliminación de los PM/NTC la superficie modificada mediante el procedimiento descrito no debe de someterse a secado. El dispositivo modificado puede utilizarse para la toma de cualquier tipo de medidas electroquímicas incluyendo, pero no exclusivamente, amperométricas, voltamétricas, impedimétricas y/o conductimétricas. In order to guarantee the reuse of the devices by eliminating the PM / NTC, the surface modified by the described procedure must not be dried. The modified device can be used for the taking of any type of electrochemical measurements including, but not exclusively, amperometric, voltammetric, impedimetric and / or conductimetric.
DESCRIPCIÓN DE LOS DIBUJOS DESCRIPTION OF THE DRAWINGS
Para complementar la descripción que se está realizando y con objeto de ayudar a una mejor comprensión de las características de la invención, de acuerdo con un ejemplo preferente de realización práctica de la misma, se acompaña como parte integrante de dicha descripción, un juego de dibujos en donde con carácter ilustrativo y no limitativo, se ha representado lo siguiente: Figura 1 . Muestra un gráfica de la corriente generada en electrodos sin modificar (bare SPE) o modificados con 5μΙ de PM y cantidades crecientes de NTC (0-7.5 μΙ NTC, 1 mg/ml). Voltametría cíclica en 250 μΜ de ferrocianuro (barras oscuras) o en PBS (corriente de fondo, barras claras). To complement the description that is being made and in order to help a better understanding of the characteristics of the invention, according to a preferred example of practical implementation thereof, a set of drawings is attached as an integral part of said description. where, for illustrative and non-limiting purposes, the following has been represented: Figure 1. It shows a graph of the current generated in unmodified electrodes (bare SPE) or modified with 5μΙ of PM and increasing amounts of NTC (0-7.5 μΙ NTC, 1 mg / ml). Cyclic voltammetry in 250 μΜ of ferrocyanide (dark bars) or in PBS (background current, light bars).
Figura 2. Muestra un gráfica de la corriente generada en los electrodos sin modificar (bare SPE) o modificados con 5μΙ de NTC y cantidades crecientes de PM (1 -10 μΙ). Voltametría cíclica en 250 μΜ de ferrocianuro (barras oscuras) o en PBS (corriente de fondo, barras claras).  Figure 2. Shows a graph of the current generated in the unmodified electrodes (bare SPE) or modified with 5μΙ of NTC and increasing amounts of PM (1 -10 μΙ). Cyclic voltammetry in 250 μΜ of ferrocyanide (dark bars) or in PBS (background current, light bars).
Figura 3. Muestra unos voltamogramas cíclicos obtenidos para concentraciones crecientes de dopamina (DA), disuelta en PBS, sobre un electrodo sin modificar. Figure 3. It shows cyclic voltamograms obtained for increasing concentrations of dopamine (DA), dissolved in PBS, on an unmodified electrode.
Figura 4. Muestra unos voltamogramas cíclicos obtenidos para concentraciones crecientes de DA, disuelta en PBS, sobre un electrodo modificado con 10 μΙ de PM y 5μΙ de NTC depositados mediante atrapamiento magnético. Figure 4. It shows cyclic voltamograms obtained for increasing concentrations of DA, dissolved in PBS, on an electrode modified with 10 μΙ of PM and 5μΙ of NTC deposited by magnetic entrapment.
Figura 5. Muestra una gráfica con los valores de corriente de pico obtenidos para concentraciones crecientes de DA en electrodos modificados y sin modificar. Figure 5. Shows a graph with the peak current values obtained for increasing concentrations of DA in modified electrodes and without Modify.
Figura 6. Muestra una gráfica de una voltametría de pulso diferencia (DPV) obtenida en dopamina (DA) y en ácido úrico (AU) medidos en un electrodo sin modificar. Figura 7. Muestra un gráfica de una voltametría de pulso diferencia (DPV) obtenida en dopamina (DA) y en ácido úrico (AU) medidos en un electrodo modificado con PM/NTC. Figure 6. Shows a graph of a difference pulse voltammetry (DPV) obtained in dopamine (DA) and uric acid (AU) measured on an unmodified electrode. Figure 7. Shows a graph of a difference pulse voltammetry (DPV) obtained in dopamine (DA) and uric acid (AU) measured on a modified electrode with PM / NTC.
REALIZACIÓN PREFERENTE DE LA INVENCIÓN PREFERRED EMBODIMENT OF THE INVENTION
A la vista de las figuras se describe a continuación un modo de realización preferente del procedimiento objeto de esta invención. In view of the figures, a preferred embodiment of the process object of this invention is described below.
Se usaron NTC de pared simple (SWNTC) modificados con grupos carboxilo (-COOH) en su superficie. Los NTC fueron resuspendidos en Solución Salina Amortiguada por Fosfatos (PBS) a una concentración final de 1 mg/ml y se dispersaron por sonicación durante al menos 30 minutos a temperatura ambiente. Seguidamente se agitó la suspensión usando un vórtex durante 2 minutos. Estos NTC son solubles en medio acuoso hasta 1 mg/ml; presentan un diámetro individual promedio de 1 ,4 nm ± 0,1 nm y forman haces de 4-5 nm 0.5- 1.5 mieras. Al principio de cada día de trabajo, y con el fin de garantizar su funcionamiento óptimo, se sonicaban los NTC durante 15 minutos, seguido de agitación al vórtex durante 1 minuto más. Las PM tenían 2.8 mieras de diámetro y estaban recubiertas de proteína.Single wall NTC (SWNTC) modified with carboxyl groups (-COOH) on their surface were used. The NTCs were resuspended in Phosphate Buffered Saline (PBS) at a final concentration of 1 mg / ml and dispersed by sonication for at least 30 minutes at room temperature. The suspension was then stirred using a vortex for 2 minutes. These NTCs are soluble in aqueous medium up to 1 mg / ml; they have an average individual diameter of 1.4 nm ± 0.1 nm and form beams of 4-5 nm 0.5-1.5 microns. At the beginning of each work day, and in order to ensure their optimal functioning, the NTCs were sonicated for 15 minutes, followed by vortexing for another 1 minute. The PM were 2.8 microns in diameter and were coated with protein.
Las MP se recibían a una concentración aproximada de E 3-3.5 x 108 partículas por mililitro (equivalente a 3-3,5 x 105 MP, aproximadamente 5 mg, por microlitro). Las MP se agitaban usando un vórtex durante un minuto, para resuspenderlas totalmente antes de su utilización, y el volumen necesario era transferido a un tubo Eppendorf. A continuación las PM eran concentradas con la ayuda de un imán o un primer generador de campo magnético, el sobrenadante era eliminado, las PM se lavaban 3 veces con PBS (10mM Phosphate Buffer Saline, pH 7.4) y se resuspendían en PBS a la concentración elegida. The MPs were received at an approximate concentration of E 3-3.5 x 10 8 particles per milliliter (equivalent to 3-3.5 x 10 5 MP, approximately 5 mg, per microliter). The MPs were agitated using a vortex for one minute, to completely resuspend them before use, and the necessary volume was transferred to an Eppendorf tube. Then the PM were concentrated with the help of a magnet or a first magnetic field generator, the supernatant was removed, PM were washed 3 times with PBS (10mM Phosphate Buffer Saline, pH 7.4) and resuspended in PBS at the chosen concentration.
Se usaron dispositivos electrónicos constituidos por electrodos serigrafiados planos integrados por un electrodo de trabajo (1.6 mm de diámetro) y un electrodo auxiliar de oro, y un electrodo de referencia de Ag/AgCI, todos ellos impresos sobre un sustrato cerámico. Previo a su utilización, los electrodos se activaron electroquímicamente mediante voltametría cíclica entre 0 y 1.4 V en 1 M H2S04 hasta que se obtuvo una señal estable. La caracterización posterior se llevó a cabo por voltametría cíclica en 0.1 M K3[Fe(CN)6]. Electronic devices consisting of flat screen-printed electrodes composed of a working electrode (1.6 mm in diameter) and a gold auxiliary electrode, and an Ag / AgCI reference electrode, all of them printed on a ceramic substrate, were used. Prior to use, the electrodes were electrochemically activated by cyclic voltammetry between 0 and 1.4 V in 1 MH 2 S0 4 until a stable signal was obtained. Subsequent characterization was carried out by cyclic voltammetry in 0.1 M K3 [Fe (CN) 6].
Para llevar a cabo el atrapamiento magnético, debajo del electrodo de trabajo se fijó un imán cilindrico de 1 mm de diámetro a modo de segundo generador de campo magnético. In order to carry out the magnetic entrapment, a 1 mm diameter cylindrical magnet was fixed under the working electrode as a second magnetic field generator.
Una vez realizada la detección o medida haciendo uso del dispositivo electrónico con el electrodo recubierto, se retira el segundo generador de campo magnético del envés del electrodo de trabajo para desprender el recubrimiento producido anteriormente sobre el electrodo de trabajo; quedando el dispositivo en su estado original. Once the detection or measurement has been carried out using the electronic device with the coated electrode, the second magnetic field generator is removed from the back of the working electrode to release the previously produced coating on the working electrode; leaving the device in its original state.
EJEMPLO 1. Eficiencia de electrodos modificados por atrapamiento magnético de cantidades crecientes de NTC. EXAMPLE 1. Efficiency of electrodes modified by magnetic entrapment of increasing amounts of NTC.
Se recubrió un electrodo, el electrodo de trabajo, con una cantidad fija de PM (1 ,5 x 106 MP), previamente incubadas con cantidades crecientes de NTC (1- 10 μg) resultando una mezcla en suspensión. Dicha mezcla se depositó sobre el electrodo de trabajo, bajo el que se había colocado el segundo generador de campo magnético. El precipitado formado por MP y NTC sobre el electrodo de trabajo, visible a simple vista, se lavó con PBS sin ser alterado. La eficiencia del electrodo modificado se estudió entonces mediante voltametría cíclica (de -0.1 a +0.45 V, a velocidad de escaneo 100 mV) en PBS (control negativo / corriente de fondo) y en ferrocianuro de potasio 250 μΜ. An electrode, the working electrode, was coated with a fixed amount of PM (1.5 x 10 6 MP), previously incubated with increasing amounts of NTC (1-10 μg) resulting in a suspension mixture. Said mixture was deposited on the working electrode, under which the second magnetic field generator had been placed. The precipitate formed by MP and NTC on the working electrode, visible to the naked eye, was washed with PBS without being altered. The efficiency of the modified electrode was then studied by cyclic voltammetry (from -0.1 to +0.45 V, at 100 mV scanning speed) in PBS (negative control / background current) and in 250 μΜ potassium ferrocyanide.
Como se resume en la Figura 1 , el atrapamiento de MP sin NTC genera pasivacion parcial del electrodo, aumento de la corriente de fondo en PBS y disminución de la corriente registrada en ferrocianuro. Por otra parte, el atrapamiento de cantidades crecientes de NTC resulta en aumento adicional de la corriente de fondo en PBS, cuyo valor está directamente relacionado con la cantidad de NTC capturados, e induce la aparición de un pre-pico entre -0,1 y 0 V. En presencia de ferricianuro, el atrapamiento de cantidades crecientes de NTC se correlaciona con un aumento lineal en la señal registrada en ferricianuro para cantidades entre 1 y 5 μg de NTC. As summarized in Figure 1, the entrapment of MP without NTC generates partial passivation of the electrode, increase in the background current in PBS and decrease in the current recorded in ferrocyanide. On the other hand, entrapment of increasing amounts of NTC results in an additional increase in the background current in PBS, whose value is directly related to the amount of NTCs captured, and induces the appearance of a pre-peak between -0.1 and 0 V. In the presence of ferricyanide, entrapment of increasing amounts of NTC correlates with a linear increase in the signal recorded in ferricyanide for amounts between 1 and 5 μg of NTC.
EJEMPLO 2. Eficiencia de electrodos modificados por atrapamiento magnético mediante cantidades crecientes de PM. EXAMPLE 2. Efficiency of electrodes modified by magnetic entrapment by increasing amounts of PM.
Se recubrió un electrodo, el electrodo de trabajo, con una cantidad fija de NTC (5 μg) mediante atrapamiento magnético usando un número variable de MP (0 - 3 x 106 MP). Los reactivos, electrodos y procedimiento experimental son similares a los descritos en el ejemplo 1. An electrode, the working electrode, was coated with a fixed amount of NTC (5 μg) by magnetic entrapment using a variable number of MP (0 - 3 x 10 6 MP). The reagents, electrodes and experimental procedure are similar to those described in example 1.
Como se observa en la Figura 2, los números de PM más bajos ensayados generaron depósitos de pequeño tamaño en la superficie del electrodo. En estos casos la mayoría de los NTC añadidos no eran atrapados y eran eliminados durante los lavados. Para los electrodos utilizados, era necesario emplear un mínimo de 2 μΙ de MP para el atrapamiento de NTC para que los electrodos modificados generaran señales consistentemente más altas que las registradas en los electrodos sin modificar. El incremento en la cantidad de MP utilizadas se correlaciona con una mayor cantidad de NTC atrapados. Esto se traduce en disminución de la cantidad de NTC eliminados durante las fases de lavado e incremento simultáneo de la corriente de fondo registrada en PBS y de la corriente de pico en ferricianuro. A partir de 10 μΙ de MP, sin embargo, se detecta saturación de la señal. Esto se corresponde con el recubrimiento total de la superficie del electrodo con PM/NTC. En estas condiciones, la voltametría en ferricianuro genera un pico un 407% más alto que el pico generado en un electrodo sin modificar. As seen in Figure 2, the lowest PM numbers tested generated small deposits on the electrode surface. In these cases, most of the added NTCs were not trapped and were removed during washing. For the electrodes used, it was necessary to use a minimum of 2 μΙ MP for NTC trapping so that the modified electrodes generated signals consistently higher than those recorded on the unmodified electrodes. The increase in the amount of MP used is correlated with a greater amount of trapped NTC. This translates into a decrease in the amount of NTC removed during the washing phases and simultaneous increase in the background current recorded in PBS and the current peak in ferricyanide. From 10 μΙ MP, however, signal saturation is detected. This corresponds to the total coating of the electrode surface with PM / NTC. Under these conditions, ferricyanide voltammetry generates a peak 407% higher than the peak generated on an unmodified electrode.
EJEMPLO 3. Detección de dopamina (DA) usando electrodos modificados por atrapamiento magnético de PM/NTC. EXAMPLE 3. Detection of dopamine (DA) using electrodes modified by magnetic entrapment of PM / NTC.
El tercer ejemplo incluye la detección de una molécula electroactiva de interés clínico, la dopamina (DA), usando electrodos modificados con PM/NTC por atrapamiento magnético. La DA es un neurotransmisor liberado por el hipotálamo y el mal funcionamiento de su actividad ha sido relacionado con varios trastornos neurodegenerativos, incluidos la anorexia y la bulimia, las enfermedades de Parkinson y de Alzheimer, y la esquizofrenia. Los reactivos, electrodos y procedimiento experimental son similares a los descritos en el ejemplo 1 . En este caso se modificó la superficie con 3 x 106 PM resuspendidas en 5μg de NTC. La DA fue disuelta en PBS a diferentes concentraciones finales y fue analizada por voltametría cíclica usando en paralelo electrodos sin modificar, tal y como se observa en la Figura 3, y electrodos modificados con PM/NTC, tal y como se observa en la Figura 4. En los electrodos sin modificar la DA genera un pico de oxidación a 0,2 V (vs. Ag/AgCI) y un pico de reducción mucho menor a -0.05 V (vs. Ag/AgCI). Como otros autores han observado anteriormente, la oxidación deThe third example includes the detection of an electroactive molecule of clinical interest, dopamine (DA), using electrodes modified with PM / NTC by magnetic entrapment. DA is a neurotransmitter released by the hypothalamus and the malfunction of its activity has been linked to several neurodegenerative disorders, including anorexia and bulimia, Parkinson's and Alzheimer's diseases, and schizophrenia. The reagents, electrodes and experimental procedure are similar to those described in example 1. In this case the surface was modified with 3 x 10 6 PM resuspended in 5μg of NTC. DA was dissolved in PBS at different final concentrations and was analyzed by cyclic voltammetry using unmodified electrodes in parallel, as seen in Figure 3, and electrodes modified with PM / NTC, as seen in Figure 4. In the electrodes without modifying the DA generates an oxidation peak at 0.2 V (vs. Ag / AgCI) and a reduction peak much less than -0.05 V (vs. Ag / AgCI). As other authors have noted previously, the oxidation of
DA es poco reversible, porque el producto oxidado se adsorbe en la superficie del electrodo. En el caso del electrodo modificado con PM/NTC, las corrientes de fondo registradas son más altas, en consonancia con el aumento de la superficie del electrodo debido a la deposición de los NTC. La separación entre picos es de 0, 10 V para el electrodo modificado, frente a 0,15 V en el electrodo sin modificar.DA is not very reversible, because the oxidized product adsorbs on the electrode surface. In the case of the modified electrode with PM / NTC, the recorded bottom currents are higher, in line with the increase in the electrode surface due to the deposition of the NTC. The separation between peaks is 0.10 V for the modified electrode, compared to 0.15 V for the unmodified electrode.
Esta significativa mejora denota la fuerte actividad catalítica de la superficie modificada hacia la DA. El incremento de la corriente de pico, por otra parte, puede atribuirse a un efecto combinado del aumento de superficie en el electrodo modificado, la acumulación de DA cargada positivamente sobre la superficie del electrodo, que gracias a los NTC está cargada negativamente, y la actividad electrocatalítica del electrodo modificado. This significant improvement denotes the strong catalytic activity of the modified surface towards AD. The increase in peak current, on the other hand, It can be attributed to a combined effect of the surface increase in the modified electrode, the accumulation of positively charged DA on the electrode surface, which thanks to the NTC is negatively charged, and the electrocatalytic activity of the modified electrode.
Para los dos tipos de electrodos (modificados y sin modificar), la corriente del pico de oxidación aumenta con la concentración de DA. Sin embargo, la reversibilidad de la reacción mejora en los electrodos modificados con NTC, y también lo hace la detectabilidad de DA. Por ejemplo, las señales registradas son, para todas las concentraciones ensayadas de DA, de 2,6 a 5 veces mayores en los electrodos modificados que las señales generadas en los electrodos sin modificar. La Figura 5 muestra el promedio de datos, calculado a partir de los resultados de tres electrodos independientes. El límite de detección, calculado como la media de al menos cinco blancos más tres veces la desviación estándar de los mismos, mejora de 1 ,69 μΜ en los electrodos sin modificar a 0,71 μΜ cuando se utilizan electrodos con PM/NTC. La sensibilidad del ensayo, medida como la pendiente de la parte lineal de la gráfica, también mejora en superficie modificada con PIW NTC (30 vs 9 nAmp/μΜ DA). For both types of electrodes (modified and unmodified), the oxidation peak current increases with the concentration of DA. However, the reversibility of the reaction improves on the electrodes modified with NTC, and so does the detectability of DA. For example, the recorded signals are, for all tested DA concentrations, 2.6 to 5 times higher in the modified electrodes than the signals generated in the unmodified electrodes. Figure 5 shows the average data, calculated from the results of three independent electrodes. The detection limit, calculated as the average of at least five targets plus three times the standard deviation thereof, improves from 1.69 μ 69 in the unmodified electrodes to 0.71 μΜ when electrodes with PM / NTC are used. The sensitivity of the test, measured as the slope of the linear part of the graph, also improves on surface modified with PIW NTC (30 vs 9 nAmp / μΜ DA).
EJEMPLO 4. Detección simultánea de dopamina y ácido úrico usando electrodos modificados por atrapamiento magnético de PM/NTC. EXAMPLE 4. Simultaneous detection of dopamine and uric acid using electrodes modified by magnetic entrapment of PM / NTC.
La principal limitación de la detección de la DA es la interferencia de componentes como el ácido úrico (AU), presentes en muestras reales a concentraciones más altas que la DA. El AU se oxida en la mayoría de los electrodos a potenciales tan cercanos a los de la DA que los picos generados por ambos compuestos son difícilmente distinguibles. La incorporación de NTC al electrodo acostumbra a mejorar la resolución de los picos generados por la DA y el AU. En este ejemplo comparamos la detección por Voltametría de Pulso Diferencial (DPV) de DA y AU, por separado o mezclados a una concentración de 200 μΜ cada uno, el los electrodos sin modificar o modificados con PM/NTC ya descritos en el ejemplo 3. Como se ilustra en las Figuras 6 y 7, DA y AU analizados por separado generan picos de oxidación mayores en los electrodos modificados con PM/NTC que en los electrodos sin modificar (9,77 vs 3,91 μΑ para la DA y 6,53 vs 1 ,11 μΑ para el AU, en electrodos modificados / sin modificar respectivamente). Este efecto es más evidente para el AU, que genera un pico mucho más claro y casi 6 veces mayor en el electrodo modificado. El potencial de pico del AU se desplaza negativamente 64 mV en los electrodos modificados, lo cual confirma la actividad catalítica de los mismos. The main limitation of the detection of AD is the interference of components such as uric acid (AU), present in real samples at concentrations higher than DA. The AU oxidizes in most electrodes to potentials so close to those of the DA that the peaks generated by both compounds are hardly distinguishable. The incorporation of NTC to the electrode usually improves the resolution of the peaks generated by the DA and the AU. In this example we compare the detection by Differential Pulse Voltammetry (DPV) of DA and AU, separately or mixed at a concentration of 200 μΜ each, the electrodes unmodified or modified with PM / NTC already described in example 3. As illustrated in Figures 6 and 7, DA and AU analyzed separately generate higher oxidation peaks in electrodes modified with PM / NTC than in unmodified electrodes (9.77 vs. 3.91 μΑ for DA and 6, 53 vs 1, 11 μΑ for AU, on modified / unmodified electrodes respectively). This effect is more evident for the AU, which generates a much clearer and almost 6 times higher peak in the modified electrode. The peak potential of the AU shifts 64 mV negatively on the modified electrodes, which confirms their catalytic activity.
La detección simultánea de DA y AU en los electrodos sin modificar evidencia un importante nivel de electrocatálisis de la DA por el AU, tal y como se observa en la figura 6. Esto se traduce en aumento de la altura del pico de la DA cuando se mide en presencia de AU a concentración equimolar, respecto a la corriente generada por una concentración similar de DA sola. Al mismo tiempo, el pico del AU disminuye en altura en presencia de DA y se vuelve casi indetectable. Por el contrario, en los electrodos modificados con PM/NTC los dos picos están claramente separados y no hay casi diferencias en la altura de los picos que se puedan atribuir a la catálisis de la DA por el AU tal y como se observa en la Figura The simultaneous detection of DA and AU in the unmodified electrodes evidences an important level of electrocatalysis of the DA by the AU, as can be seen in Figure 6. This translates into an increase in the height of the DA peak when measured in the presence of AU at equimolar concentration, with respect to the current generated by a similar concentration of DA alone. At the same time, the AU peak decreases in height in the presence of DA and becomes almost undetectable. On the contrary, in the electrodes modified with PM / NTC the two peaks are clearly separated and there are almost no differences in the height of the peaks that can be attributed to the catalysis of the DA by the AU as seen in the Figure

Claims

R E I V I N D I C A C I O N E S
1 . Procedimiento de recubrimiento de electrodos de un dispositivo electrónico que comprende: a) activar y/o lavar un electrodo de trabajo a recubrir, one . Electrode coating method of an electronic device comprising: a) activating and / or washing a working electrode to be coated,
b) dispersar unos nanotubos de carbono (NTC) en un medio acuoso a una concentración definida entre 0.1 y 10 mg/ml  b) disperse carbon nanotubes (NTC) in an aqueous medium at a defined concentration between 0.1 and 10 mg / ml
c) concentrar unas partículas magnéticas (PM) haciendo uso de un campo magnético generado por un primer generador de campos magnéticos, d) eliminar sobrenadante, e) lavar las partículas magnéticas (PM) con el medio acuoso en que se han dispersado los NTC y repetir c) y d),  c) concentrate magnetic particles (PM) using a magnetic field generated by a first generator of magnetic fields, d) remove supernatant, e) wash the magnetic particles (PM) with the aqueous medium in which the NTCs have been dispersed and repeat c) and d),
f) resuspender el resultado de la fase anterior en un volumen de dispersión de NTC preparado en b) g) fijar un segundo generador de campo magnético en el envés del electrodo de trabajo a recubrir,  f) resuspend the result of the previous phase in a volume of NTC dispersion prepared in b) g) fix a second magnetic field generator on the underside of the working electrode to be coated,
h) recubrir el electrodo de trabajo por deposición del resultado de (f) sobre una cara a recubrir del electrodo de trabajo.  h) coating the working electrode by deposition of the result of (f) on a face to be coated of the working electrode.
2. Procedimiento según reivindicación 1 caracterizado porque incluye retirar el generador de campo magnético causando el desprendimiento del recubrimiento presente sobre el electrodo de trabajo y lavar el electrodo de trabajo.  2. Method according to claim 1 characterized in that it includes removing the magnetic field generator causing the detachment of the coating present on the working electrode and washing the working electrode.
3. -Procedimiento según reivindicación 1 caracterizado porque los NTC son solubles en medio acuoso. 3. Procedure according to claim 1 characterized in that the NTCs are soluble in aqueous medium.
4. Procedimiento según reivindicación 1 caracterizado porque las PM son solubles en medio acuoso. 4. Method according to claim 1 characterized in that the PMs are soluble in aqueous medium.
5. - Procedimiento según reivindicación 1 caracterizado porque las PM están recubiertas por al menos una proteína. 5. - Method according to claim 1 characterized in that the PMs are coated by at least one protein.
6. - Procedimiento según reivindicación 1 caracterizado porque el medio acuoso de b) es una solución salina. 6. - Method according to claim 1 characterized in that the aqueous medium of b) is a saline solution.
7. Procedimiento según cualquiera de las reivindicaciones anteriores caracterizado porque los NTC están modificados.  7. Method according to any of the preceding claims characterized in that the NTCs are modified.
8. Procedimiento según reivindicación 6 caracterizado porque los NTC están funcionalizados. 8. Method according to claim 6 characterized in that the NTCs are functionalized.
9. Electrodo obtenible mediante el procedimiento descrito en una cualquiera de las reivindicaciones 1 a 8.  9. Electrode obtainable by the method described in any one of claims 1 to 8.
10. Dispositivo electrónico caracterizado porque comprende el electrodo descrito en la reivindicación 9.  10. Electronic device characterized in that it comprises the electrode described in claim 9.
1 1 . Uso del dispositivo electrónico descrito en la reivindicación anterior para la realización de detección y medición de analitos.  eleven . Use of the electronic device described in the preceding claim for the detection and measurement of analytes.
PCT/ES2010/070853 2009-12-23 2010-12-21 Method for coating electrodes of an electronic device by means of magnetic trapping, resulting electrode, device including same and use of said device WO2011086209A1 (en)

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Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2004052489A2 (en) * 2002-12-09 2004-06-24 The University Of North Carolina At Chapel Hill Methods for assembly and sorting of nanostructure-containing materials and related articles
US20070275627A1 (en) * 2006-05-26 2007-11-29 Korea Advanced Institute Of Science And Technology Method for fabricating field emitter electrode using array of carbon nanotubes
WO2008066965A2 (en) * 2006-06-23 2008-06-05 The Regents Of The University Of California Articles comprising large-surface-area bio-compatible materials and methods for making and using them

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2004052489A2 (en) * 2002-12-09 2004-06-24 The University Of North Carolina At Chapel Hill Methods for assembly and sorting of nanostructure-containing materials and related articles
US20070275627A1 (en) * 2006-05-26 2007-11-29 Korea Advanced Institute Of Science And Technology Method for fabricating field emitter electrode using array of carbon nanotubes
WO2008066965A2 (en) * 2006-06-23 2008-06-05 The Regents Of The University Of California Articles comprising large-surface-area bio-compatible materials and methods for making and using them

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