DK2576830T3 - Fremgangsmåder til in situ detektering af nukleotidsekvenser - Google Patents
Fremgangsmåder til in situ detektering af nukleotidsekvenser Download PDFInfo
- Publication number
- DK2576830T3 DK2576830T3 DK11731575.4T DK11731575T DK2576830T3 DK 2576830 T3 DK2576830 T3 DK 2576830T3 DK 11731575 T DK11731575 T DK 11731575T DK 2576830 T3 DK2576830 T3 DK 2576830T3
- Authority
- DK
- Denmark
- Prior art keywords
- approx
- sample
- nucleic acid
- target nucleic
- oligonucleotide probe
- Prior art date
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Classifications
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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
- C12Q1/6841—In situ hybridisation
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- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Life Sciences & Earth Sciences (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Health & Medical Sciences (AREA)
- Engineering & Computer Science (AREA)
- Microbiology (AREA)
- Immunology (AREA)
- Physics & Mathematics (AREA)
- Molecular Biology (AREA)
- Biotechnology (AREA)
- Biophysics (AREA)
- Analytical Chemistry (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)
Claims (25)
1. Fremgangsmåde til at bestemme hvorvidt en target-nukleinsyre er til stede i en biologisk prøve på en fast bærer, omfattende trinnene at: a) at bringe prøven på bæreren i kontakt med en denatureringsbuffer omfattende en base og ca. 50 til ca. 80% alkohol, hvor basen er til stede i opløsningen ved en koncentration på ca. 0,03N til ca. 0,17N, derved denaturere prøven; b) inkubere den denaturerede prøve i en hybridiseringsbuffer der omfatter mindst én enkelt-strenget oligonukleotid-probe ved en temperatur i området på ca. 19°C til ca. 25°C, hvor oligonukleotid-proben omfatter en nukleotidsekvens der er i det væsentlige komplementær med en nukleotidsekvens i target-nukleinsyren og mindst én detekterbar markør; c) vaske prøven for at fjerne ikke-hybridiserede oligonukleotid-prober; og d) bestemme hvorvidt target-nukleinsyren er til stede i prøven ved at detektere én eller flere oligonukleotid-prober der har hybridiseret til target-nukleinsyren i prøven.
2. Fremgangsmåde ifølge krav 1, hvor basen er natriumhydroxid.
3. Fremgangsmåde ifølge krav 1 eller 2, hvor trin a) udføres ved en temperatur på ca. 19°C til ca. 25°C, fx ved en temperatur på ca. 21°C.
4. Fremgangsmåde ifølge krav 3, hvor prøven bringes i kontakt med opløsningen i ca. 3 til ca. 20 minutter i trin a), fx i ca. 11 til ca. 17 minutter i trin a), og fortrinsvis i ca. 13 til ca. 15 minutter i trin a).
5. Fremgangsmåde ifølge krav 3, hvor opløsningen i trin a) omfatter ca. 0,07M natriumhydroxid.
6. Fremgangsmåde ifølge krav 5, hvor opløsningen i trin a) omfatter ca. 70% ethanol.
7. Fremgangsmåde ifølge krav 1 eller 2, hvor trin b) udføres ved en temperatur på ca. 21°C.
8. Fremgangsmåde ifølge krav 1, hvor, før trin b), oligonukleotid-proben er i en hybridiseringsbuffer der omfatter formamid, dextransulfat, og ét eller flere salte ved en slutkoncentration på ca. 0,03M til ca. 0,09M.
9. Fremgangsmåde ifølge krav 1, hvor trinnet til at fjerne ikke-hybridiserede oligonukleotid-prober fra prøven omfatter vaskning af prøven i en vaskebuffer ved en temperatur på ca. 19°C til ca. 25°C før trin c).
10. Fremgangsmåde ifølge krav 9, hvor vaskebufferen omfatter ét eller flere salte ved en slutkoncentration på ca. 0,03M til ca. 0,09M, og natriumdodecylsulfat (SDS).
11. Fremgangsmåde ifølge krav 1, hvor oligonukleotid-proben omfatter ca. 20 til ca. 50 nukleotider, fx ca. 30 nukleotider.
12. Fremgangsmåde ifølge krav 11, hvor oligonukleotid-proben er en syntetisk oligonukleotid-probe.
13. Fremgangsmåde ifølge krav 1, hvor den mindst ene detekterbare markør er bundet til oligonukleotidet ved en kovalent binding.
14. Fremgangsmåde ifølge krav 13, hvor den mindst ene detekterbare markør omfatter en fluorescerende markør.
15. Fremgangsmåde til atdetektere en target-nukleinsyre i en biologisk prøve på en fast bærer, omfattende trinnene at: a) at bringe prøven på bæreren i kontakt med en denatureringsbuffer omfattende en base og ca. 50 til ca. 80% alkohol, hvor basen er til stede i opløsningen ved en koncentration på ca. 0,03N til ca. 0,17N, derved denaturere prøven; b) hybridisere mindst en enkelt-strenget oligonukleotid-probe til target-nukleinsyren i den denaturerede prøve i en hybridiseringsbuffer ved en temperatur i området på ca. 19°C til ca. 25°C, hvor oligonukleotid-proben omfatter en nukleotidsekvens der er i det væsentlige komplementær med en nukleotidsekvens i target-nukleinsyren og mindst én detekterbar markør; c) vaske prøven for at fjerne ikke-hybridiserede oligonukleotid-prober; og d) detektere den mindst ene detekterbare markør på oligonukleotid-proben efterfulgt af hybridisering til target-nukleinsyren i prøven, derved detektere target-nukleinsyren i prøven.
16. Fremgangsmåde ifølge krav 15, hvor basen er natriumhydroxid.
17. Fremgangsmåde ifølge krav 1 eller 15, hvor den biologiske prøve omfatter urothelial-celler.
18. Kit til at detektere en target-nukleinsyre i en biologisk prøve, omfattende: a) mindst én enkelt-strenget oligonukleotid-probe bestående af ca. 20 til ca. 50 nukleotider, hvor mindst én detekterbar markør er kovalent bundet til oligonukleotid-proben; b) denatureringsbuffer omfattende ca. 0,03M til ca. 0,17M natriumhydroxid og ca. 50 til ca. 80% alkohol; c) hybridiseringsbuffer omfattende ca. 20 til ca. 90% formamid, dextransulfat, og ét eller flere salte ved en slutkoncentration på ca. 0,03M til ca. 0,09M; og d) en vaskebuffer der omfatter ét eller flere salte ved en slutkoncentration på ca. 0,03M til ca. 0,09M, og ca. 0,1 % SDS.
19. Kittet ifølge krav 18, hvor denatureringsbufferen omfatter ca. 0,07M natriumhydroxid og ca. 70% ethanol.
20. Kittet ifølge krav 18 eller 19, hvor hybridiseringsbufferen omfatter ca. 60 til ca. 80% formamid.
21. Kittet ifølge krav 18, hvor det ene eller flere salte i vaskebufferen er valgt fra gruppen bestående af et natriumsalt, et lithiumsalt og et kaliumsalt.
22. Kittet ifølge krav 18, hvor det ene eller flere salte i vaskebufferen omfatter natriumcitrat og natriumchlorid.
23. Kittet ifølge krav 21 eller 22, hvor vaskebufferen yderligere omfatter formamid.
24.
Fremgangsmåde til at detektere en target-nukleinsyre i en biologisk prøve på en fast bærer, omfattende trinnene at: a) at bringe prøven på bæreren i kontakt med en denatureringsbuffer omfattende ca. 0,07M natriumhydroxid and ca. 70% ethanol for ca. 13 til ca. 15 minutter ved en temperatur i området på ca. 19°C til ca. 25°C, derved denaturere prøven; b) hybridisere mindst en enkelt-strenget oligonukleotid-probe bestående af ca. 20 til ca. 50 nukleotider til target-nukleinsyren i den denaturerede prøve i en hybridiseringsbuffer ved en temperatur i området på ca. 19°C til ca. 25°C, hvor oligonukleotid-proben omfatter en nukleotidsekvens der er i det væsentlige komplementær med en nukleotidsekvens i target-nukleinsyren, og mindst én fluorescerende detekterbar markør kovalent bundet til oligonukleotid-proben; c) vaske prøven for at fjerne ikke-hybridiserede oligonukleotid-prober; og d) detektere den fluorescerende detekterbar markør på oligonukleotid-proben, derved detektere target-nukleinsyren i prøven.
Applications Claiming Priority (2)
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|---|---|---|---|
| US35106410P | 2010-06-03 | 2010-06-03 | |
| PCT/US2011/038934 WO2011153354A1 (en) | 2010-06-03 | 2011-06-02 | Methods and kits for in situ detection of nucleotide sequences |
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| Publication Number | Publication Date |
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| DK2576830T3 true DK2576830T3 (da) | 2014-12-08 |
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| EP (1) | EP2576830B1 (da) |
| BR (1) | BR112012030813A2 (da) |
| DK (1) | DK2576830T3 (da) |
| WO (1) | WO2011153354A1 (da) |
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| JP5588970B2 (ja) | 2008-05-27 | 2014-09-10 | ダコ・デンマーク・エー／エス | ハイブリダイゼーション組成物及び方法 |
| US9303287B2 (en) | 2009-02-26 | 2016-04-05 | Dako Denmark A/S | Compositions and methods for RNA hybridization applications |
| US10662465B2 (en) | 2011-09-30 | 2020-05-26 | Agilent Technologies, Inc. | Hybridization compositions and methods using formamide |
| US11118226B2 (en) | 2011-10-21 | 2021-09-14 | Agilent Technologies, Inc. | Hybridization compositions and methods |
| US9145584B2 (en) | 2011-12-12 | 2015-09-29 | Cellay, Inc. | Methods and kits for room temperature in situ detection of a target nucleic acid in a biological sample |
| BR112016010354B1 (pt) | 2013-11-07 | 2019-09-17 | Cellay, Inc. | Métodos e kits para detectar um carcinoma de próstata e prever prognósticos de doença para cânceres de próstata |
| US10525076B2 (en) | 2015-02-20 | 2020-01-07 | Rosalind Franklin University Of Medicine And Science | Antisense compounds targeting genes associated with cystic fibrosis |
| EP3259356B1 (en) * | 2015-02-20 | 2021-12-01 | Rosalind Franklin University of Medicine and Science | Antisense compounds targeting genes associated with cystic fibrosis |
| WO2017041005A1 (en) | 2015-09-03 | 2017-03-09 | Abbott Molecular Inc. | Hybridization buffers comprising an alkyl diester |
| US10457981B2 (en) | 2016-01-08 | 2019-10-29 | Abbott Molecular Inc. | Hybridization buffers |
| EP4556575A3 (en) | 2016-11-21 | 2025-10-08 | Bruker Spatial Biology, Inc. | A method for sequencing nucleic acids |
| EP3794146B1 (en) | 2018-05-14 | 2025-12-10 | Bruker Spatial Biology, Inc. | Method for identifying a predetermined nucleotide sequence |
| JP2023521356A (ja) * | 2020-04-07 | 2023-05-24 | アドヴァンスド セル ダイアグノスティクス，インコーポレイテッド | ｉｎ ｓｉｔｕハイブリダイゼーションによって標的核酸を検出する方法及びそのキット |
| CN113063646B (zh) * | 2021-03-27 | 2022-06-21 | 复旦大学 | 一种细胞固定剂、细胞固定方法和应用 |
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| US5753437A (en) * | 1987-10-13 | 1998-05-19 | The United States Of America As Represented By The Department Of Health & Human Services | Method of diagnosing cancer susceptibility or metastatic potential |
| US5082830A (en) | 1988-02-26 | 1992-01-21 | Enzo Biochem, Inc. | End labeled nucleotide probe |
| NL9001639A (nl) | 1990-07-19 | 1992-02-17 | Amc Amsterdam | Pt-houdende verbinding, werkwijze voor de bereiding ervan, alsmede toepassing van dergelijke verbindingen. |
| US5714327A (en) | 1990-07-19 | 1998-02-03 | Kreatech Diagnostics | Platinum-containing compounds, methods for their preparation and applications thereof |
| US5719024A (en) | 1993-08-18 | 1998-02-17 | Applied Spectral Imaging Ltd. | Method for chromosome classification by decorrelation statistical analysis and hardware therefore |
| EP1019420B1 (en) | 1995-05-09 | 2003-08-06 | Kreatech Biotechnology B.V. | Methods for the production of platinum-based linkers between labels and bio-organic molecules, for labelling bio-organic molecules, for detecting biological substances of interest |
| US6780603B1 (en) * | 1995-07-21 | 2004-08-24 | Regents Of The University Of Minnesota | Analysis of alpha integrins for the diagnosis of diabetic nephropathy |
| CZ114199A3 (cs) | 1996-10-04 | 1999-09-15 | Novo Nordisk A/S | N-Substituované azaheterocyklické sloučeniny |
| US20020197656A1 (en) * | 1999-12-17 | 2002-12-26 | Ronghao Li | Cell arrays and the uses thereof |
| US20040115692A1 (en) * | 2000-04-03 | 2004-06-17 | Cytyc Corporation | Methods, compositions and apparatuses for detecting a target in a preservative solution |
| US6617148B2 (en) * | 2000-06-29 | 2003-09-09 | E. I. Du Pont De Nemours And Company | Natural promoters for gene expression and metabolic monitoring in bacillus species |
| ATE334230T1 (de) | 2001-03-02 | 2006-08-15 | Stratagene California | Zusammensetzungen und verfahren mit platinverbindungen zur nukleinsäuremarkierung |
| US20040158883A1 (en) * | 2002-11-05 | 2004-08-12 | Crawford Fiona C. | Organic anion transport polypeptide related protein-4 (OATPRP4) gene in Tourette syndrome and related disorders |
| DE602005022714D1 (de) * | 2004-06-03 | 2010-09-16 | Advandx Inc | Hybridisierung von pna-sonden in alkohollösungen |
| US20090036664A1 (en) * | 2007-07-31 | 2009-02-05 | Brian Jon Peter | Complex oligonucleotide primer mix |
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2011
- 2011-06-02 WO PCT/US2011/038934 patent/WO2011153354A1/en not_active Ceased
- 2011-06-02 DK DK11731575.4T patent/DK2576830T3/da active
- 2011-06-02 EP EP11731575.4A patent/EP2576830B1/en not_active Not-in-force
- 2011-06-02 BR BR112012030813A patent/BR112012030813A2/pt not_active IP Right Cessation
- 2011-06-02 US US13/701,310 patent/US20130203055A1/en not_active Abandoned
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| BR112012030813A2 (pt) | 2017-06-20 |
| WO2011153354A1 (en) | 2011-12-08 |
| US20130203055A1 (en) | 2013-08-08 |
| WO2011153354A8 (en) | 2013-01-03 |
| EP2576830A1 (en) | 2013-04-10 |
| EP2576830B1 (en) | 2014-08-27 |
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