WO2016190727A1 - Biocapteur électrochimique d'adn pour l'identification du sexe et de la variété - Google Patents

Biocapteur électrochimique d'adn pour l'identification du sexe et de la variété Download PDF

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WO2016190727A1
WO2016190727A1 PCT/MY2016/000031 MY2016000031W WO2016190727A1 WO 2016190727 A1 WO2016190727 A1 WO 2016190727A1 MY 2016000031 W MY2016000031 W MY 2016000031W WO 2016190727 A1 WO2016190727 A1 WO 2016190727A1
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dna
biosensor
electrochemical
dna biosensor
fish
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PCT/MY2016/000031
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English (en)
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Yook Heng Lee
Abd Rashid ZULKAFLI
Poh Chiang CHEW
Futra DEDI
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Universiti Kebangsaan Malaysia
Jabatan Perikanan Malaysia
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Priority to CN201680043018.7A priority Critical patent/CN107922978A/zh
Publication of WO2016190727A1 publication Critical patent/WO2016190727A1/fr

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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6813—Hybridisation assays
    • C12Q1/6816—Hybridisation assays characterised by the detection means
    • 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
    • G01N27/3276—Sensing specific biomolecules, e.g. nucleic acid strands, based on an electrode surface reaction being a hybridisation with immobilised receptors
    • 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

  • Embodiments of the present invention relate to a biosensor and more particularly to an electrochemical DNA biosensor which rapidly recognizes the sample DNA for identifying the gender and variety of Arowana fishes without the need of any skillful operator. Further, it can be miniaturized according to the needs of usage. Also, it provides an advantage of portability and user friendly structure.
  • Conventional biosensor consists of a bio-recognition component, a bio-transducer component, and an electronic system which include a signal amplifier, a processor, and a display.
  • the bio-recognition component interacts with analyteof interest and the interaction is measured by the bio-transducer which outputs a measurable signal proportional to the presence of the target analyte in the sample.
  • the analyte detected can be both organic and inorganic in nature.
  • WO 2008099163 A1 describes a method of detection of the protein-dependent coincidence of DNA in a sample which comprises detection using luminescence of one or more luminophores introduced into DNAwith one or more DNA fragments in which fragments are bound using one or more DNA-binding proteins. Further, fluorescence technique comprises the use of ALEX-FRET.
  • US20060228738 A1 describes a DNA-polypyrrole based biosensor and methods of using the biosensor for the rapid detection of Escherichia Coli and other microorganisms.
  • the DNA-polypyrrole biosensor is used to detect micoorganisms for monitoring water quality of a sample from a drinking water or food source. Further, the biosensor uses genomic DNA extracted from natural environments for the rapid detection of microorganisms to provide an early warning of water contamination.
  • CN 103529036A describes a sex identification method for juvenile fish of hybrid Pelteobagrusfulvidraco (Richardson).
  • the hybrid fish is identified by a method of living-body gonad squashing, acetocarmine staining and ethanol crystal violet staining. Further, the accuracy of the method is verified by a paraffin section of the gonad tissue.
  • the biosensor can detect trace amount of sample solution.
  • the sample to be analyzed is exposed to additional chemicals that attach to the analytes in order to tag or mark them.
  • the tagged analytes become much bigger and respond differently to specific frequencies of light or otherwise change physical properties in some way which makes them easier to detect.
  • conventionalbiosensorscan provide various advantages, such as those described above, conventional biosensorsare limited in various other matters such asuse of radioactively-labeled or redox-labeled probes is problematic as the radioactive labels are short-lived which require the completion of analysis within a short period of time. Further, the use of redox-labels depends on color change that may suffer interference in real samples.
  • analyte detection systems are based on analyte-specific binding between an analyte and an analyte-binding receptor.
  • Such systems typically require complex multicomponent detection systems such as Enzyme-Linked Immunosorbent Assay (ELISA), electrochemical detection systems or require that both the analyte and the receptor are labeled with detection molecules for example Fluorescence Resonance Energy Transfer (FRET) systems.
  • ELISA Enzyme-Linked Immunosorbent Assay
  • FRET Fluorescence Resonance Energy Transfer
  • FRET Fluorescence Resonance Energy Transfer
  • PCR Polymerase Chain Reaction
  • gel electrophoresis which can be done only by a skillful operator. Further, these techniques are too slowand labour intensive and can be performed only in laboratories.
  • an electrochemical DNA biosensor which rapidly recognizes a sample DNA for identifying the gender and variety of Arowana fishes via electrochemical detection. Further, the biosensor is portable and provides simple and rapid results. Also, it can be operated by unskilled operators such as farmers.
  • Embodiments of the present invention aim to provide an electrochemical DNA biosensor which rapidly recognizes a sample DNA for identifying gender and variety of Arowana fishes via electrochemical detection. Further, the proposed biosensor measures the current produced which does not depend on color change and thus eliminates the risk of interference in real samples. Furthermore, the electrochemical DNA biosensor involvesdirect detection method and gives rapid results both qualitatively and quantitatively without the need of PCR amplification reaction. Also, it is user-friendly and portable.
  • the present invention is provided with the features of claim 1 , however the invention may additionally reside in any combination of features of claim 1.
  • the electrochemical DNA biosensor comprising a screen-printed electrode modified with gold nanoparticles, a layer of silica nanosphere composite, an oligonucleotide sequence, a cross-linking agent and a redox indicator.
  • the layer of silica nanosphere composite isdepositedon the screen-printed electrode and the oligonucleotide sequence is immobilized on the layer of silica nanosphere composite using the cross-linking agent.
  • the oligonucleotide sequence is complementary to a unique target sequence of a nucleic acid from a fish of interest such that the nucleic acid from the fish of interest hybridizes to the oligonucleotide sequence to form a dsDNA.
  • the redox mediator intercalates with the dsDNA which enables detection of gender and variety of the fish by voltammetry.
  • the screen-printed electrode is a carbon screen-printed electrode.
  • oligonucleotide sequence is a DNA sequence of a Malaysian Golden Arowana (Scleropagesformosus) fish.
  • the cross-linking agent is glutaricdialdehyde (GA).
  • the redox indicator is antraquinonemonosulphonic acid (AQMS).
  • the target sequence is a DNA or RNA.
  • the fish is Malaysian Golden Arowana (Scleropagesformosus).
  • the electrochemical DNA biosensorfurther comprising a dry reagent pad including buffer salt and redox indicator. Further, the redox indicator and buffer salt are immobilized in the dry reagent pad.
  • the voltammetry is differential pulse voltammetry (DPV).
  • DUV differential pulse voltammetry
  • compositions or an element or a group of elements are preceded with the transitional phrase "comprising”, it is understood that we also contemplate the same composition, element or group of elements with transitional phrases “consisting of, “consisting”, “selected from the group of consisting of, “including”, or “is” preceding the recitation of the composition, element or group of elements and vice versa.
  • Fig. 1 illustrates an exploded view of anelectrochemical DNA biosensor electrode in accordance with an embodiment of the present invention.
  • Fig. 2 is a graph showing differential pulse voltammograms of AQMS current from the electrochemical DNA biosensor by using SiNSp modified gold nanoparticles-SPE electrode.
  • Fig. 3 is a graph showing effect of gold nanoparticles (A), silica nanospheres (B), DNA probe concentrations (C), temperature (D), DNA hybridization times (E) and pH of Na-phosphate buffer (F) on the electrochemical DNA biosensor response verified by AQMS indicator.
  • Fig. 4 is a graph showing differential pulse voltamograms (A) and calibration curves (B) of DPV peak current response generated by the electrochemical DNA biosensor.
  • Fig. 5 is a graph showing regeneration performance of the electrochemical DNA biosensor with NaOH (0.1 M) as a regeneration solution (15 min) (B) and re-hybridization of DNA probe immobilized on Si-Au-SPE with 1.0 x10 "9 M cDNA (30 min) (A).
  • the present invention is described hereinafter by various embodiments with reference to the accompanying drawing, wherein reference numerals used in the accompanying drawing correspond to the like elements throughout the description. This invention may, however, be embodied in many different forms and should not be construed as limited to the embodiment set forth herein.
  • the present invention is able to provide an alternative electrochemical DNA biosensor to replace the conventional biosensors.
  • the disclosed electrochemical DNA biosensor is reagentless. In other words, no addition of reagent is possible during its operation.
  • the proposed electrochemical DNA biosensor measures the current produced in order to detect the sample. Also, it is portable and provides rapid results on site (for example, within 15 mins) without the need of any skillful operator.
  • the electrochemical DNA biosensor comprising a screen-printed electrode modified with gold nanoparticles, a layer of silica nanosphere composite, an oligonucleotide sequence, a cross-linking agent and a redox indicator.
  • the layer of silica nanosphere composite is deposited on the screen-printed electrode and the oligonucleotide sequence is immobilized on the layer of silica nanosphere composite using the cross-linking agent.
  • the oligonucleotide sequence is complementary to a unique target sequence of a nucleic acid from a fish of interest such that the nucleic acid from the fish of interest hybridizes to the oligonucleotide sequence to form a dsDNA.
  • the redox indicator intercalates with the dsDNA which enables detection of gender and variety of the fish by voltammetry.
  • the screen-printed electrode is fabricated from a composite of silica nanospheres (SiNSp) and gold nanoparticles (AuNPs), to form a modified carbon screen-printed electrode (Si-Au-SPE).
  • the silica nanospheres (SiNSp) are biocompatible and facilitates high DNA loading. Also, they provide large surface area and ease of depositing on the electrode.
  • the oligonucleotide sequence (DNA probe)is covalently coupled onto the silica nanosphere layer through glutaricdialdehyde (GA)which acts as a cross linking agent.
  • GA glutaricdialdehyde
  • the oligonucletode sequence is complementary to the target DNA sequence of Arowana fish.
  • the target DNA sequence is a cDNA extracted from the Arowana fish samples.
  • target cDNA hybridizes with the oligonucleotide sequence (DNA probe) to form a dsDNA.
  • the redox indicator, anthraquainone-2-sulfonic acid monohydrate sodium (AQMS) salt intercalates with the dsDNA and enables the detection of the gender and variety of Malaysian Golden Arowana fish using differential pulse voltammetry (DPV).
  • DPV differential pulse voltammetry
  • the electrochemical DNA biosensor includes a dry reagent pad.
  • the dry reagent pad includesredox indicator, buffer salts and polyvinyl alcohol (PVA).Further, theredox indicator and buffer salts are immobilized in the dry reagent pad to avoid the use of chemicals.
  • PVA polyvinyl alcohol
  • the dry reagent pad allows a slow release of the redox indicators as they are intercalated with the dsDNA.
  • the oligonucleotide sequence for both gender and variety determination involved multi-probes, that is 2-3 different probes will be involved for either gender or variety determination. These probes were obtained from molecular techniques (PCR, cloning and electrophoresis) on more than a hundred samples of Arowana fish tissues. Two to three probes with the best specificity to the Arowana fish gender or variety were selected and later used for biosensor fabrication. These probes are of 20- 25bp in length.
  • a three-electrode system of electrochemical DNA biosensor electrode (100),as shown in Figure 1 was utilized for the DNA biosensor measurement with carbon paste screen-printed electrode (Scrint Technology (M)) as a working electrode (102), a printed Ag/AgCI electrode as a reference electrode(106) and a carbon paste counter electrode (104).
  • the DNA biosensor includes a dry reagent pad (108) which further includes buffer salt and redox indicator immobilized in polyvinyl alcohol (PVA). Upon hybridization the dry reagent pad (108) allows a slow release of redox indicator.
  • the screen-printed electrodes are transducer for current when the target cDNA hybridization occurs with the DNA probe.
  • Electrochemical measurements were performed using differential pulse voltammetry (DPV) with anAutolab PGSTAT 12 potentiostat (Metrohm, Ultrecht, Netherlands).
  • the parameters for DPV was 0.02 V step potential in the scan range -0.85 to -0. 5 V.
  • Amino functionalized silica nanospheres were produced by using the modified Tang et al method. A mixture of deionized water (2 mL), ammonia solution (5 mL) and ethanol (20 mL) was sonicated for 10 min at room temperature. Thereafter, a mixture of tetraethoxysilane (TEOS) (2 mL) and ethanol (4 mL) was added to above mentioned mixture and sonicated for another 30 min at 56°C. The silica nanospheres colloid was directly functionalized with 3-aminopropyltriethoxysilane (APTS) (2 ml) and stirred overnight at 25°C.
  • TEOS tetraethoxysilane
  • APTS 3-aminopropyltriethoxysilane
  • AuNPs gold nanoparticles
  • Au-SPE gold nanoparticles
  • silica nanospheres colloid 2 ⁇ _.
  • the Si-Au-SPE was soaked in 400 ⁇ _ glutaricdialdehyde (GA) (5 %) for two hours to activateaminatedSiNPs' surface and rinsed with deionized water.
  • the GA functionalized Si-Au-SPE was incubated in 300 ⁇ _ DNA probe (2 ⁇ ) solution overnight at 4°C and rinsed carefully with K-phosphate buffer (0.05 M, pH 7.0) to remove the unbound DNA probe.
  • the immobilized DNA probe was immersed in 300 ⁇ _ target cDNA solution containing NaCI (1 M) and AQMS (1 mM) for 30 min to allow hybridization process and rinsed thoroughly with deionised water several times and immersed in K-phosphate buffer for 6 min to release physically adsorbed cDNA and/or AQMS. All measurement of the DPV current signal was carried out in 450 ⁇ _ of K- phosphate buffer (0.05 M, pH 7.0) at 25°C. Optimization of DNA biosensor response
  • the optimization of DNA biosensor response was observed to obtain the best working condition of the biosensor.
  • Gold nanoparticles were loaded from 0.01 to 0.05 mg .
  • Silica nanospheres stacking was optimized between 0.001 to 0.05 mg onto the Au-SPE.
  • the enhanced DNA biosensor response was evaluated by immobilizing the DNA probe on the silica nanospheres modified Au-SPE electrode between 0.01 to 30 ⁇ . Further, the DNA hybridization reaction was incubated in water bath from 16°C to 60°C. Whilst optimum hybridization time of the DNA biosensor was evaluated by immersing the DNA probe immobilized Si-Au-SPE into the complementary DNA (cDNA) solution from 1 to 240 min. To determine optimum H, the pH of Na-phosphate buffer was verified from pH 3.0 to pH 9.0.
  • the buffer salt and PVA concentration in the dry reagent pad were verified from 0.01 - 0.5 M and 1-10%, respectively.
  • the amount of AQMS immobilized in PVA was also optimized.
  • the leaching time was optimized from 15 - 120 min. DNA hybridization determination
  • the DNA biosensors were investigated in a series concentration of cDNA target solution from 1.0 * 10 "19 to 1.0 * 10 "7 M.
  • the DNA biosensor signal was collected after 30 min DNA hybridization reaction at 25°C. While reproducibility of the DNA biosensors were batch prepared by using two different cDNA concentrations i.e. 1.0 * 10 "11 and 1.0 * 10 "9 M.
  • the DPV signals of 10 units of DNA biosensor was evaluated after 30 min hybridization reaction. Regeneration of the proposed DNA biosensor was performed by incubating the immobilized DNA probe in 300 ⁇ _ of DNA target solution (1.0 * 10 "9 M) for 30 min at 25°C.
  • the DNA biosensor was soaked on 0.1 M of NaOH solution for 15 min to introduce dsDNA breaks and washed carefully with K-phospate buffer (0.05 M, pH 7.0) for 2 min.
  • K-phospate buffer 0.05 M, pH 7.0
  • the deposited DNA probe was carried out hybridization with 1.0 ⁇ 10 "9 M cDNA again for 30 min at room temperature.
  • the extracted DNA concentration used for determination of Arowana variety was 100 fold dilutions using Na-phosphate buffer (0.05 M, pH 7.0). About 300 ml_ DNA extraction solution containing NaCI (1 M) and AQMS (1 mM) was sonicated for 15 min to release the dsDNA breaks. Then, the immobilized DNA probe was soaked for 30 min to allow DNA hybridization process and washed carefully with deionized water to remove unbound or adsorbed extracted DNA. Evaluation of Arowana DNA variety based on the DPV peak current signal was measured after DNA hybridization reaction and compared with DPV current response of control. Whereas control was DNA probe functionalized Si-Au-SPE electrode without analyte (extracted Arowana DNA).
  • AuNPs gold nanoparticles
  • SiNSp silica nanospheres
  • DNA probe temperature
  • DNA hybridization time pH of Na-phosphate buffer
  • pH of Na-phosphate buffer pH of Na-phosphate buffer
  • the DNA biosensor signal increased as the AuNPs depositing increased from 0.001 to 0.003 mg ( Figure3A) due to the increasing electron transfer from AQMS redox mediator to the gold nanoparticles modified SPE electrode surface.
  • the AuNPs quantities increased from 0.003 to 0.01 mg, the DNA biosensor response decreased because the increasing
  • the DNA biosensor response increased with the SiNSp loading from 0.002 to 0.014 mg (Figure 3B). This indicated that increased SiNSp concentration could largely prepare active amine site of SiNPs to react with DNA probe followed by hybridization reaction with cDNA which further increase AQMS intercalation with dsDNA.
  • the SiNSp concentration increased from 0.014 to 0.02 mg
  • the DNA biosensor response reduced as the higher amount of the SiNSp has blocked the electron transfer from AQMS redox to the SPE electrode surface.
  • optimum AuNPs and SiNSp loading was 0.003 mg and 0.012 mg, respectively.
  • the DNA biosensor signal gradually increased with DNA probe solution loading from 0.1 to 2.0 ⁇ . This implicated that the increasing DNA probe quantities were reacted with the active amine site of SiNSp resulting in increased DNA hybridization and AQMS intercalation with dsDNA.
  • the DNA biosensor response was sustained with DNA probe solution loading from 2.0 to 3.0 ⁇ , which related that the DNA probe were fully coupled with cDNAs.
  • the DNA biosensor response increased steadily from 5 to 30 min of DNA hybridization time (Figure 3E) with DNA probe as the high amount of cDNA interacted with the DNA probe on the Si-Au-SPE electrode surface.
  • the DNA biosensor signal was found to be constant when the DNA hybridization reaction time continued from 45 to 240 min as the immobilized DNA probe on the Si-A-SPE electrode had been fully coupled with target cDNA.
  • the optimum DNA hybridization time was selected after 30 min keeping DNA probe in the cDNA solution.
  • Na-phosphate buffer which results in the increasing electrostatic repulsion between negatively charged phosphodiester DNA molecules.
  • Na-phosphate buffer at pH 6.5 was chosen as the optimum pH for the DNA hybridization reaction.
  • the optimum concentration of buffer salt and PVA for the dry reagent pad was found to be 0.05 M and 5%, respectively.
  • the optimum leaching time was found out to be 30 min.
  • the optimum amount of AQMS immobilized in PVA in the dry reagent pad was found to be 0.715 mg.
  • the calibration curves of the DNA biosensor were observed by using various targets Arowana DNA concentrations from 1.0 * 10 "13 -1.0 ⁇ 10 "1 ⁇ with 30 min DNA hybridization at 25°Care shown in Figure 4.
  • the DNA hybridization response increased proportionally with increasing amount of cDNA immobilized on the silica nanospheres modified Au-SPE electrode due to the increasing DNA hybridization and AQMS intercalation in the dsDNA on the electrode surface ( Figure 4A).
  • the Si-Au composites based on DNA biosensor showed that the linear response range was in the range of 1.0 * 10 "17 to 1.0 * 10 "7 M and the lowest detection limit was 1.4 x 10 ⁇ 18 M.
  • the reproducibility of DNA biosensor was evaluated by using 10 different electrodes and performed with two cDNA concentration i.e.
  • the regeneration performance of the developed biosensor response is shown in Figure 5.
  • the DNA biosensor response declined after dipping in NaOH solution (0.1 M) for 15 min as the hydrogen bonding between base pairs of dsDNA wasbroken by NaOH solution and OH " ions interacted with hydrogen atom from sugar phosphate backbone of DNA followed by dsDNA breaks.
  • DNA biosensor limit to accuracy is 1-2 bp mismatch. In other words, it cannot detect ⁇ 2 bp mismatch.
  • the DNA biosensor is able to detect different strains of Arowanawhen a specific DNA probe is used.
  • the electrochemical DNAbiosensor has commercial potential in pathogen detection in aquaculture, agriculture and human disease diagnosis.
  • the electrochemical DNA biosensor is useful as it is fast, sensitive and is of great utility for monitoring food safety and quality, particularly by monitoring microbial contamination and environmental contamination by biohazards.
  • electrochemical DNA biosensor as described above could be fabricated in various other ways and could include various other materials, including various other DNA probes, electrodes, salts etc.

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Abstract

L'invention concerne un biocapteur électrochimique d'ADN, comprenant une électrode imprimée par sérigraphie modifiée par des nanoparticules d'or, une couche de composite de nanosphères de silice, une séquence oligonucléotidique, un agent de réticulation et un indicateur redox. La couche de composite de nanosphères de silice est déposée sur l'électrode imprimée par sérigraphie et la séquence oligonucléotidique est immobilisée sur la couche de composite de nanosphères de silice à l'aide de l'agent de réticulation. En outre, au moins une partie de la séquence oligonucléotidique est complémentaire à une séquence cible unique d'un acide nucléique d'un poisson d'intérêt, de telle sorte que l'acide nucléique du poisson d'intérêt s'hybride à la séquence oligonucléotidique pour former un ADN à double brin. En outre, l'indicateur redox s'intercale avec l'ADN à double brin, ce qui permet la détection du sexe et de la variété du poisson par voltamétrie.
PCT/MY2016/000031 2015-05-22 2016-05-23 Biocapteur électrochimique d'adn pour l'identification du sexe et de la variété WO2016190727A1 (fr)

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WO2018223024A3 (fr) * 2017-06-01 2019-03-07 The Regents Of The University Of California Mesure sans étalonnage avec biocapteurs électrochimiques
WO2020176792A1 (fr) * 2019-02-27 2020-09-03 California Institute Of Technology Approche de détection électrochimique pour la quantification de molécules dans des fluides corporels

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US20130334063A1 (en) * 2012-06-15 2013-12-19 Gordon & Rosenblatt, Llc Method of Detecting Analyte
CN103760201A (zh) * 2013-12-10 2014-04-30 天津工业大学 一种基于复合量子点的电化学dna传感器的制备方法

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WO2005108625A2 (fr) * 2001-07-13 2005-11-17 Nanosphere, Inc. Procede pour la preparation de substrats comportant des molecules immobilisees et substrats
WO2010117341A1 (fr) * 2009-04-08 2010-10-14 Agency For Science, Technology And Research Biocapteur d'acide nucléique
US20130334063A1 (en) * 2012-06-15 2013-12-19 Gordon & Rosenblatt, Llc Method of Detecting Analyte
CN103760201A (zh) * 2013-12-10 2014-04-30 天津工业大学 一种基于复合量子点的电化学dna传感器的制备方法

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Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2018223024A3 (fr) * 2017-06-01 2019-03-07 The Regents Of The University Of California Mesure sans étalonnage avec biocapteurs électrochimiques
US11946098B2 (en) 2017-06-01 2024-04-02 The Regents Of The University Of California Calibration-free measurement with electrochemical biosensors
WO2020176792A1 (fr) * 2019-02-27 2020-09-03 California Institute Of Technology Approche de détection électrochimique pour la quantification de molécules dans des fluides corporels
US11549934B2 (en) 2019-02-27 2023-01-10 California Institute Of Technology Electrochemical sensing approach for molecule quantification in body fluids

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