EP1456417A2 - Procede de determination d'analytes du type acide nucleique - Google Patents

Procede de determination d'analytes du type acide nucleique

Info

Publication number
EP1456417A2
EP1456417A2 EP02805308A EP02805308A EP1456417A2 EP 1456417 A2 EP1456417 A2 EP 1456417A2 EP 02805308 A EP02805308 A EP 02805308A EP 02805308 A EP02805308 A EP 02805308A EP 1456417 A2 EP1456417 A2 EP 1456417A2
Authority
EP
European Patent Office
Prior art keywords
nucleic acid
sensor
sensors
analyte
solid phase
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Withdrawn
Application number
EP02805308A
Other languages
German (de)
English (en)
Inventor
Holger Klapproth
Mirko Lehmann
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
TDK Micronas GmbH
Original Assignee
TDK Micronas GmbH
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by TDK Micronas GmbH filed Critical TDK Micronas GmbH
Publication of EP1456417A2 publication Critical patent/EP1456417A2/fr
Withdrawn legal-status Critical Current

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Classifications

    • 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
    • C12Q1/6825—Nucleic acid detection involving sensors
    • 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
    • C12Q1/6834—Enzymatic or biochemical coupling of nucleic acids to a solid phase
    • C12Q1/6837—Enzymatic or biochemical coupling of nucleic acids to a solid phase using probe arrays or probe chips
    • 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
    • G01N27/403—Cells and electrode assemblies
    • G01N27/414—Ion-sensitive or chemical field-effect transistors, i.e. ISFETS or CHEMFETS

Definitions

  • the present invention relates generally to a method and a device for determining nucleic acid analyzes.
  • the invention relates to the detection of the presence of such an analyte without the conventional use of optically detectable marker substances.
  • biosensors or biochips For the qualitative and / or quantitative determination of certain nucleic acid analytes such as DNA, the use of essentially planar systems is known, which are referred to in the art as biosensors or biochips. These biochips form a carrier, on the surface of which a plurality of detection regions, which are usually arranged in a grid-like manner, are usually formed, the individual regions or region groups each differing from one another in their specificity with respect to a specific analyte to be detected.
  • nucleic acid probes such as oligonucleotides or cDNA in mostly single-stranded form
  • sequence sequence probe design
  • the chip surface that is functionalized in this way is brought into contact with the DNA analytes to be determined as part of a corresponding detection method under conditions which, if the target nucleic acid (s) previously detected are present, ensure their hybridization with the immobilized probe molecules.
  • the qualitative and, if necessary, quantitative detection of one or more specifically formed hybridization complexes is then usually carried out by optophysical luminescence measurement and assignment of the data obtained to the respective detection areas, which enables the determination, for example, of the presence or the sequence of the nucleic acid analyzer (s) and, if necessary, their quantification.
  • Another approach of the prior art relates to the utilization of the enzymatic activity of the extracellular endonuclease of Serra tia marcescens, the enzyme-mediated degradation of DNA leading to a change in the pH, which can then be measured with a pH sensor (S. Reher , DNA, RNA analysis with voltametric, potentiometric and optical methods using the extracellular endonuclease Serra tia marcescens, ISBN 3- 89825-030-X, 1999).
  • a pH sensor S. Reher , DNA, RNA analysis with voltametric, potentiometric and optical methods using the extracellular endonuclease Serra tia marcescens, ISBN 3- 89825-030-X, 1999.
  • DNA analysis is described using certain marker substances, but the detection is not carried out by optical methods.
  • the object of the present invention is therefore to provide an improved method in which the disadvantages associated with the drift problem are overcome.
  • the object is achieved by the method according to the main claim.
  • the present invention relates to a method for determining a nucleic acid analyte by hybridizing the analyte to a suitable nucleic acid probe immobilized on a solid phase, in which
  • the analyte is determined on the basis of physical measurement data which are specifically related to an enzyme-related mass increase or decrease in the hybridization complex, the measurement of the data being carried out by at least one sensor which is an integral part of the solid phase.
  • This indirect approach enables the detection to be transferred to another time window and thus to a different frequency, which preferably comprises only a few seconds or minutes (see FIG. 1) while largely avoiding the drift problem.
  • determination used in the present context refers to any analysis of a nucleic acid and includes in particular the detection of the presence of a nucleic acid analyte in a sample to be investigated.
  • forms of use such as the determination of a nucleic acid sequence and the detection of mutations such as, in particular, SNPs are also included Procedure guaranteed thus a very wide range of possible uses, since it is based on all currently and future available determination or Detection techniques are applicable, which are based on the formation of a hybridization complex.
  • the enzyme causing the mass increase or decrease of the hybridization complex is selected from the group consisting of polymerases, ligases, ribozymes, quasi-catalytic nucleic acids, DNases / RNAses (exo- or endonucleases including restriction endonucleases), and RNase H, wherein a polymerase, in particular a polymerase having a 5 ⁇ - and / or 3 x -Exonukleasemodtician, is particularly preferred.
  • RNA-dependent DNA polymerases a mass increase depending on the given nature of the nucleic acid (RNA or DNA) can also take place according to the invention by using RNA-dependent DNA polymerases (reverse transcriptase) or RNA-dependent RNA polymerases (replicases) .
  • a mass increase can also be brought about by using appropriate polymerase-active ribozymes or quasi-catalytic RNAs.
  • polymerase-active ribozymes or quasi-catalytic RNAs for all polymerases (including ribozymes or quasi-catalytic RNAs) it is the invention that both thermostable and thermolabile enzymes can be used.
  • An enzymatically caused mass increase can also take place by means of ligases.
  • ligases including ribozymes or quasi-catalytic RNAs.
  • thermostable and thermolabile enzymes like can be used for all ligases (including ribozymes or quasi-catalytic RNA 's).
  • thermostable and thermolabile enzymes like can be used for all ligases (including ribozymes or quasi-catalytic RNA 's).
  • thermostable and thermolabile enzymes like can be used.
  • a decrease in mass can also be detected.
  • a decrease in mass by cleavage of the bound nucleic acids can be done by nucleases (RNases, DNases). Both 5 - and / or 3 ⁇ - exo- as well as endonucleases and RNase H can be used.
  • Ribozymes or quasi-catalytic RNAs with nuclease activity are also suitable. Ribozymes or quasi-catalytic RNAs generally act in a sequence-specific manner, the specificity being able to be adjusted according to the needs by means of the respective hybridization sequence.
  • the present invention accordingly reflects the situation found in most solid-phase-bound nucleic acid analyzes of the presence of a single-stranded nucleic acid probe immobilized on a solid phase (see FIG. 1A). Under suitable conditions, an at least partially at least double-stranded hybridization complex forms on this probe in the presence of a nucleic acid analyte which is essentially complementary to the probe sequence (see FIG. 1B).
  • an enzymatic step is initiated (see FIG. IC), the enzyme performance leading to a measurable change in the mass of this complex.
  • the polymerase activity mentioned above gives rise to pyrophosphate anions which are released in the polymerization of the nucleotide triphosphates in the single-stranded region of the hybridization complex and lead to local acidification and thus to a lowering of the pH.
  • a pH sensor or a pH detector e.g. pH-ISFET
  • this change in the pH value can be detected, if desired location-specifically (see Fig. 2).
  • the pyrophosphate ions released during a polymerization or ligation can also be detected indirectly, ie via a secondary enzyme cascade.
  • a secondary enzyme cascade For example, ATP-sulfurylase and adenosine 5'-phosphosulfate (APS) are involved in the first of the secondary reactions.
  • the pyrophosphate PPi released during the incorporation of a nucleotide during the polymerization or during a ligation and the APS are converted to ATP by the ATP sulfurylase.
  • the ATP generated in this way can then catalyze further enzymatic reactions which are fed to the actual detection.
  • the ATP formed, the reaction of luciferin by the luciferase catalyze arises whereby a light emission which 'can be scanned with the inventive optical sensors.
  • a modified example of the method according to the invention relates to the loading of the nucleotide triphosphates to be used with magnetic beads (Baselt, loc. Cit., 1998) or with metal particles (Clinical micro sensors, loc. Cit., 2000). The result of this loading is that the reading is further enhanced by additional properties of the solid body bound to the nucleotide triphosphate.
  • dyes can be present on the nucleotide triphosphate, which can then be read out via an integrated photodiode.
  • the at least one sensor is therefore selected from the group consisting of electrode structures, field effect transistors, magnetic sensors, optical sensors and pH sensors in order to correspond to the wide range of applications of the present invention.
  • a particularly preferred embodiment relates to the combinatorial use of different sensors of the aforementioned type.
  • the signal intensity or sharpness and thus the reliability of a desired detection event could be optimized if a device suitable for the method according to the invention not only a probe-specific sensor, e.g. a field effect transistor, but also another type of probe-specific sensor, e.g. has a pH-ISFET.
  • the data obtained from this multiparametric measurement enable an even more precise evaluation of the enzyme-related signals.
  • the senor can be equipped with a heating element.
  • a heating element could consist, for example, of conductor tracks which are applied during the CMOS process and subsequently by the following Layers have been covered. In this way, temperature cycles could be carried out, which could be desirable, for example, for a PCR-supported application in the context of the method according to the invention.
  • an apparatus for performing the method according to the invention is provided.
  • This device comprises at least one solid phase, at least one nucleic acid probe immobilized directly or indirectly thereon, and at least one sensor for recording the physical measurement data, the sensor being an integral part of the solid phase and preferably being selected from the previously defined group consisting of electrode structures and field effect transistors , Magnetic sensors, optical sensors and pH sensors.
  • a large number of different nucleic acid probes are arranged in a grid-like manner with the formation of a microarray, with each immobilized nucleic acid probe or each specific detection area being particularly preferably assigned at least one sensor.
  • the measuring device known from EP-A-0 881 490 for measuring certain physiological as well as morphological parameters of at least one living cell to be examined can be used accordingly for the use according to the invention Modification can be used.
  • the device described already has a large number of sensors which are an integral part of a carrier device on which the material to be examined is immobilized.
  • the carrier unit of the device according to the invention essentially consists of a semiconductor material with an integrated detector layer, preferably comprising several detectors, with at least one of the sensors described above, optionally in combination, as detectors
  • the signal processing takes place at least partially within the sensor chip or chips used.
  • the sampled measurement data can be evaluated, for example, directly on the chip using analog circuits, for example by takes a value every millisecond, which then e.g. is also compared with a reference value of a previously performed measurement, which was also stored on the chip.
  • analog circuits for example by takes a value every millisecond, which then e.g. is also compared with a reference value of a previously performed measurement, which was also stored on the chip.
  • unspecific interference signals such as e.g. interfering with external signals.
  • the measurement field or point signal values can be detected sequentially, for example by detecting entire rows or columns of the sensor surface or parts thereof in succession (multiplex application ).
  • the electronic output signals of the detectors can be supplied to an external evaluation device by means of suitable circuit devices after an analog-digital conversion (see above).
  • the sensor chip surfaces such as those made of silicon dioxide
  • the sensor chip surfaces are dissolved in a solution of bifunctional molecules (so-called "linkers") which have, for example, a halosilane (for example chlorosilane) or Al oxysilane group for coupling to the carrier surface. immersed so that a self-organizing monolayer (SAM) is formed, through which the covalent bond between the sensor surface and the receptor is created.
  • linkers bifunctional molecules
  • halosilane for example chlorosilane
  • Al oxysilane group for coupling to the carrier surface.
  • glycidyltriethoxysilane can be coated, which can be done, for example, by immersion in a solution of 1% silane in toluene, slow extraction and immobilization by “baking” at 120 ° C.
  • a coating created in this way generally has a thickness of a few angstroms
  • the linker and receptor molecule (s) are coupled via a suitable further functional group, for example an amino or epoxy group, and suitable bifunctional linkers for coupling a large number of different receptor molecules, in particular also of biological origin, to a large number of support surfaces are well known to the person skilled in the art, see for example "Bioconjugate Techniques" by GT Hermanson, Academic Press 1996.
  • WO 00/43539 With regard to the formation of thin polymer layers as a coupling matrix for creating a functionalized surface, reference is made to WO 00/43539.
  • the nucleic acids provided according to the invention as probe molecules can then be applied and immobilized using conventional pressure equipment.
  • hybridizations with e.g. DNA can be performed. This can e.g. generated by PCR. When hybridizing, the DNA analyte now binds to the counter strand of the probe (if present) on the sensor. Positive hybridization events can now be detected using the method according to the invention.
  • the measurement of a location-specific mass increase can e.g. done by physical methods.
  • the location-specific change in the refractive index, the location-specific change in the electrical resistance or the electrical conductivity, the location-specific. Change in optical density or location-specific dichroic effects, etc. can be measured.
  • the general method according to the invention is suitable for a broad spectrum of fields of application, on the one hand between pure diagnostic detection of certain analytes in a sample to be analyzed and more complex derived modifications of the method for determining sequence data or information about functional ones. Connections can be distinguished in the context of corresponding genomics questions. However, this distinction is only for illustration and is in no way intended to limit the basically open applicability of the method according to the invention.
  • the method according to the invention is particularly suitable for the determination of DNA sequences which were generated by means of parallel amplification by nested (PCR), preferably in a combined liquid-phase / solid-phase DNA microarray system, since in this way both the use of modified nucleotides and eg biotin, digoxiginin, and conventionally used fluorescent dyes and other marker substances can be dispensed with.
  • PCR nested
  • the nested PCR in the combined liquid-phase / solid-phase DNA microarray system has the same sensitivity as a conventional, ie carried out in the liquid phase, PCR, but at the same time ensures a higher specificity than conventional hybridization assays and Primer extension assays.
  • the signals from the sensor are recorded by a recording unit.
  • a recording unit has a very fast converter for converting analog detector signals into digital values that are stored.
  • the digital values are preferably evaluated in real time, but can also be delayed.
  • a conventional microprocessor can be used to evaluate the digital values.
  • FIG. 1 shows the schematic sequence of an embodiment of the method according to the invention.
  • the nucleic acid probe (2) is covalently bound to the surface.
  • B After addition of the nucleic acid analyte (1), a hybridization complex is formed, usually over a period of several hours.
  • C By using a suitable enzyme, e.g. a polymerase, the single-stranded area of the complex is filled in the presence of the four nucleotides A, T, G and C (in the case of DNA) within a much shorter period of only a few minutes (D), which generates a signal much more quickly , which is read out by a sensor (4) as an integrated component of the solid phase.
  • a suitable enzyme e.g. a polymerase
  • FIG. 2 shows the principle of a preferred embodiment using a polymerase reaction, by means of which pyrophosphate anions (5) are released, which lead to a local change in the pH. This change can be detected by the integrated sensor (4).
  • FIG. 3 shows a field effect transistor produced as part of a CMOS process.
  • the field effect transistor consists, for example, of a pnp layer (6) in an n-well with a thin insulator (10) (eg 10 nm thermal oxide), which is located on the surface and to which the nucleic acid probe is applied directly or indirectly, which is then applied the hybridization comes into being.
  • the scratch protection (7) in the area of the field effect transistor is either sharp-edged or etched down in stages, so that the process of hybridization and the increase in mass (8) in a deepened area takes place.
  • the surfaces of the device can actively or passively influence the hybridization of the nucleic acid molecules by applying, for example, noble metal or hydrophobic / hydrophilic materials (9).
  • a measuring solution (11) such as 1 M NaHC0 3
  • the change in the dielectric properties on the gate is measured, which takes place by filling the single-stranded area of the hybridization complex.
  • the resulting shift in the flat band potential can be read out with the field effect transistor using a reference electrode (12) located in the solution.
  • the current between drain and source or the voltage between reference electrode and source can be recorded as a signal (see e.g. Palan et al., Fundamental Noise Limits of ISFET-Based Microsystems, poster article 4P26, • EUROSENSORS XIII (ISBN 90-76699 -02-X), pp. 169 ff., 1999).
  • FIG. 4 shows the voltage change sensed in the course of a parallel amplification on the basis of a so-called “nested on chip” PCR (NOC, see above) with an FET.
  • B the voltage curve of a single cycle (average number of cycles) in addition to the primer the template and from left to right the elongation of a primer. It can be seen from the illustration that the voltage increases as a function of the primer extension.
  • CMOS sensor is built on 5 or 6 '' wafers with a 1.2 ⁇ m
  • CMOS process manufactured. Each field effect transistor is located in an n-well on a p-type substrate. After the field oxidation, the drain and source regions are implanted. The thermal gate oxide with a thickness of approx. 10 nm is applied. The gate is protected by polysilicon during the following process steps. Then a silicon
  • the gate insulator is exposed to etching steps.
  • CMOS sensor Coating the CMOS sensor
  • the CMOS sensor prepared as above is coated with the silane by immersion in a solution of 1% GOPS (glycidoxypropyltriethoxysilane) and 0.1% triethylamine in toluene for a period of about 2 hours.
  • the chip is then removed from the solution and, after a short drop at 120 ° C., is fixed in the drying cabinet for a period of about 2 hours.
  • the chip coated in this way can be stored under exclusion of moisture until bioconjugation.
  • Bioconjugation with oligonucleotide probes Using conventional techniques, the chip coated as above is printed contact-free with 5'-amino-modified oligonucleotide probes.
  • the oligonucleotide probes are provided in a concentration of 5 ⁇ M dissolved in PBS buffer. After printing, the coupling reaction is continued at 50 ° C in a humid chamber. The chips are then rinsed with distilled water and then washed with methanol to dry. Any remaining solvent residues are finally removed by evaporation under the fume cupboard.
  • the reaction mix contains the following standard reagents (primer: 0.5 ⁇ M, dATP, dCTP, dGTP: 0, lmM, dTTP 0.08 mM, PCR buffer, MgCl 2 : 4mM, HotStarTaq (Perkin Elmer) 2 units (50 ⁇ l)
  • primer 0.5 ⁇ M, dATP, dCTP, dGTP: 0, lmM, dTTP 0.08 mM
  • PCR buffer MgCl 2 : 4mM
  • HotStarTaq Perkin Elmer 2 units
  • the above reaction mixture is in a buffer 5 x SSPE, 0.1% SDS (12 ⁇ l) under a cover slip for a period of 2 hours at 50 ° C. in the moist chamber on the chip hybridized. Then it is rinsed with 2 x SSPE 0.1% SDS and the chip is cleaned by washing in water.

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  • Chemical & Material Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Organic Chemistry (AREA)
  • Zoology (AREA)
  • Wood Science & Technology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Health & Medical Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Analytical Chemistry (AREA)
  • Microbiology (AREA)
  • Immunology (AREA)
  • Molecular Biology (AREA)
  • Biotechnology (AREA)
  • Biophysics (AREA)
  • Physics & Mathematics (AREA)
  • Biochemistry (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • General Engineering & Computer Science (AREA)
  • General Health & Medical Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)

Abstract

La présente invention concerne en général un procédé ainsi qu'un dispositif pour la détermination d'analytes du type acide nucléique. L'invention concerne en particulier la détection de la présence d'un tel analyte sans l'utilisation habituelle de substances de marquage détectables optiquement.
EP02805308A 2001-12-21 2002-12-06 Procede de determination d'analytes du type acide nucleique Withdrawn EP1456417A2 (fr)

Applications Claiming Priority (3)

Application Number Priority Date Filing Date Title
DE10163599 2001-12-21
DE10163599A DE10163599B4 (de) 2001-12-21 2001-12-21 Verfahren zur Bestimmung von Nukleinsäureanalyten
PCT/EP2002/013860 WO2003054225A2 (fr) 2001-12-21 2002-12-06 Procede de determination d'analytes du type acide nucleique

Publications (1)

Publication Number Publication Date
EP1456417A2 true EP1456417A2 (fr) 2004-09-15

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EP02805308A Withdrawn EP1456417A2 (fr) 2001-12-21 2002-12-06 Procede de determination d'analytes du type acide nucleique

Country Status (5)

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EP (1) EP1456417A2 (fr)
AU (1) AU2002356632A1 (fr)
DE (1) DE10163599B4 (fr)
TW (1) TWI310406B (fr)
WO (1) WO2003054225A2 (fr)

Families Citing this family (17)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP4832759B2 (ja) 2002-12-11 2011-12-07 サントル ナショナル ドゥ ラ ルシェルシュ シアンティフィク プローブ分子と標的生体分子との間の少なくとも１つの特異的相互作用を電子的に検出する方法
DE10309349B4 (de) * 2003-03-03 2005-11-10 Micronas Holding Gmbh Vorrichtung zur Untersuchung eines Analyten
JP3903183B2 (ja) * 2004-02-03 2007-04-11 独立行政法人物質・材料研究機構 遺伝子検出電界効果デバイスおよびこれを用いた遺伝子多型解析方法
US20050233366A1 (en) * 2004-04-16 2005-10-20 Norihisa Mino Sample-analyzing device and process for manufacturing the same
US20080138801A1 (en) * 2004-05-28 2008-06-12 Arizona Board Of Regents, A Body Corporate Acting On Behalf Of Arizona State University Surface Plasmon Resonance Sensor for Detecting Changes in Polynucleotide Mass
US7879764B2 (en) 2004-12-28 2011-02-01 Intel Corporation Electrically active combinatorial chemical (EACC) chip for biochemical analyte detection
US9695472B2 (en) * 2005-03-04 2017-07-04 Intel Corporation Sensor arrays and nucleic acid sequencing applications
US9040237B2 (en) * 2005-03-04 2015-05-26 Intel Corporation Sensor arrays and nucleic acid sequencing applications
EP2251435B1 (fr) * 2007-03-02 2013-10-16 DNA Electronics Ltd Appareil de mesure pour suivre l'amplification des acides nucléiques, utilisant un transistor à effet de champ sensible aux ions (ISFET) pour determiner le pH.
US20100047792A1 (en) * 2007-04-10 2010-02-25 Szendroe Istvan Label-free optical detection method
US8500979B2 (en) 2009-12-31 2013-08-06 Intel Corporation Nanogap chemical and biochemical sensors
US8715932B2 (en) 2010-08-20 2014-05-06 Intel Corporation Nucleic acid sequencing
US8444835B2 (en) 2010-09-09 2013-05-21 Intel Corporation Electronic and fluidic interface
US9322798B2 (en) 2011-12-15 2016-04-26 Intel Corporation Diamond electrode nanogap transducers
WO2013095635A1 (fr) * 2011-12-23 2013-06-27 Intel Corporation Instruction pour fusionner des modèles de masque
US9500617B2 (en) 2011-12-28 2016-11-22 Intel Corporation Nanogap transducers with selective surface immobilization sites
CN106872541B (zh) * 2015-12-10 2019-10-01 中山和芯生物技术有限公司 一种用于生物传感器信号放大的探针体系及其应用

Family Cites Families (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1997016699A1 (fr) * 1995-10-30 1997-05-09 Trustees Of Boston University Appareil et procedes de detection de force piezoelectrique destines au sequençage de biopolymeres
AUPO427996A0 (en) * 1996-12-20 1997-01-23 Co-Operative Research Centre For Diagnostic Technologies Method for detecting a nucleotide at a specific location within a polynucleotide sequence and apparatus therefor
JP4169827B2 (ja) * 1997-05-28 2008-10-22 ミクロナス ゲーエムベーハー 測定装置
US6287776B1 (en) * 1998-02-02 2001-09-11 Signature Bioscience, Inc. Method for detecting and classifying nucleic acid hybridization
FR2805826B1 (fr) * 2000-03-01 2002-09-20 Nucleica Nouvelles puces a adn
WO2002020832A1 (fr) * 2000-09-04 2002-03-14 Atonomics Aps Microcapteurs et procede de detection d'analytes cibles

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
See references of WO03054225A2 *

Also Published As

Publication number Publication date
DE10163599B4 (de) 2006-06-29
WO2003054225A3 (fr) 2004-04-08
TWI310406B (en) 2009-06-01
WO2003054225A2 (fr) 2003-07-03
DE10163599A1 (de) 2003-07-17
TW200301307A (en) 2003-07-01
AU2002356632A1 (en) 2003-07-09

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