DK3171172T3 - Fremgangsmåde og indretning til påvisning af bakterier - Google Patents
Fremgangsmåde og indretning til påvisning af bakterier Download PDFInfo
- Publication number
- DK3171172T3 DK3171172T3 DK15003307.4T DK15003307T DK3171172T3 DK 3171172 T3 DK3171172 T3 DK 3171172T3 DK 15003307 T DK15003307 T DK 15003307T DK 3171172 T3 DK3171172 T3 DK 3171172T3
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- DK
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- Prior art keywords
- bacteriophages
- filter
- detected
- species
- detection
- Prior art date
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Claims (15)
- PAT E NT K RAV1. Fremgangsmåde til påvisning af bakterier, omfattende trinene: A) tilvejebringelse af en suspension (1), der omfatter mindst en species af markerede testbakteriofager (2), der binder specifikt til en bakterieart, der skal påvises, hvor markeringen er kendetegnende for de respektive species af testbakteriofager (2); B) tilsætning af en prøve (3), der skal testes for forekomsten af mindst en bakterieart, der skal påvises, til suspensionen; C) blanding af prøven med suspensionen og inkubering af reaktionsblandingen; D) filtrering af reaktionsblandingen gennem et filter (4), hvor filteret (4) har en porestørrelse på 0,1 pm til 0,5 pm, og porestørrelsen af det anvendte filter (4) er valgt således, at den mindst ene bakterieart, der skal påvises, samt bakterie-bakteriofagkomplekser (5), der består af bakterier af den mindst ene bakterieart, der skal påvises, og dertil bundne testbakteriofager (2) af den mindst ene species, tilbageholdes og immobiliseres på filteroverfladen, mens ubundne bakteriofager (2, 6) kan passere filteret; E) detektion af bakterie-bakteriofagkomplekser (5) i supernatanten ved forekomst af mindst en bakterieart, der skal påvises, hvor komplekserne består af bakterier af den mindst ene bakterieart, der skal påvises, og dertil bundne testbakteriofager (2) af den mindst ene species af testbakteriofager (2); F) detektion af ubundne testbakteriofager (2) i filtratet; G) processorstøttet behandling af modtagne signaler, der frembringes ved detektionen i trinene E) og F), og udlæsning af detektionsresultaterne til en bruger, kendetegnet ved, at suspensionen endvidere omfatter en markeret reference-bakteriofag (6), der ikke binder til nogen bakterieart, der skal påvises, og en af testbakteriofagerne (2) udviser en disjunkt markering, således at den markerede referencebakteriofag (6) ligeledes detekteres i trinnet F); eller at der som alternativ til en markeret referencebakteriofag (6) anvendes markerede mikro- eller nano-partikler, der ikke binder til nogen bakterieart, der skal påvises, og på grund af deres lille størrelse kan passere filteret (4).
- 2. Fremgangsmåde til påvisning af bakterier ifølge krav 1, hvor bakteriearterne, der skal påvises, udvælges blandt gruppen omfattende Staphylococcus aureus, MethiciHin-resistent Staphylococcus aureus (MRSA), Staphylocossus epidermidis, Streptococcus pneumoniae, Streptococcus pyogenes, Haemophilus influenzae, Haemophilus parainfluenzae, Klebsiella pneumoniae, Escherichia coli, Escherichia hermannii, EHEC, Enterobacter cloacae, Enterococcus faecalis, Enterococcus faecum, Pseudomonas aeruginosa, Proteus mirabilis, Proteus vulgaris, Staphylococcus saprophyticus, Bacteroides fragilis, Entercoccus faecium, Enterobacter aerogenes, Serratia marcescens, B-Streptokokker (agalactiae), Chlamydia trachomatis, Chlamydia psittaci, Ureaplasma urealyticum, Mycoplasma hominis, Mycoplasma pneumoniae, Neisseria gonorrhoeae, Neisseria meningitidis, Citrobacter freundii, Moraxella catarrhalis, Stenotrophomonas maltophilia, Pasteurella multocida, Acinetobacter baumannii, Providencia rettgeri, Bordetella pertussis, Bacillus anthracis, Bacillus cereus, Brucella abortus, Brucella melitensis, Clostridium butolinum, Clostridium difficile, Clostridium tetani, Clostridium perfringens, Clamydia trachomatis, Corynebacterium diphtheriae, Francisella tularensis, Gardenella vaginalis, Listeria monocytogenes, Morganella morganii, Mycobacterium leprae, Mycobacterium tuberculosis, Nocardia asteriodes, Salmonella bongori, Salmonella enterica, Shigella spp., Vibrio cholerae, Borrelia burgdorferi, Yersinia pestis, Yersinia enterocolitica, Coxiella burnettii, Aeromonas spp., Plesiomonas spp., Xanthomonas maltophilia, Treponema pallidum, Eikenella corrodens, Spirillum minus, Rickettsia prowazeki, Rickettsia rickettsii, Rickettsia conorii, Cronobacter spp., Campylobacter spp. og Legionella pneumophilia.
- 3. Fremgangsmåde til påvisning af bakterier ifølge krav 1 eller 2, hvor suspensionen (1) omfatter to eller flere species af markerede testbakteriofager (2), der hver især binder specifikt til forskellige bakteriearter, der skal påvises, og hvor markeringerne af species er disjunkt.
- 4. Fremgangsmåde til påvisning af bakterier ifølge et af de foregående krav, hvor markeringen omfatter mindst et reportermolekyle med fluorescerende eller kemiluminiscerende egenskaber eller mindst et reportermolekyle, der emitterer lys ved interaktion med et sekundært molekyle.
- 5. Fremgangsmåde til påvisning af bakterier ifølge krav 4, hvor markeringen i hvert enkelt tilfælde består af mindst et fluorescerende reportermolekyle, og detektionen sker ved måling af fluorescensen, idet exciteringen af mindst et fluorescerende reportermolekyle sker i bølgelængdeområdet på 200-1000 nm.
- 6. Fremgangsmåde til påvisning af bakterier ifølge krav 4 eller 5, hvor det mindst ene reportermolekyle er koblet til proteiner i bakteriofagernes capsidregion.
- 7. Fremgangsmåde til påvisning af bakterier ifølge et af kravene 4 til 6, hvor detektionen i hvert enkelt tilfælde sker ved tidsopløst måling af lysemissionen.
- 8. Fremgangsmåde til påvisning af bakterier ifølge et af de foregående krav, der endvidere omfatter trinnet med en forseparering af bakterie-bakteriofag-komplekser (5) og ubundne bakteriofager (2, 6), hvor forsepareringen støder op til trin C) og sker ifølge princippet med gelfiltrering gennem en gelmatrix (24).
- 9. Reaktionsbeholder (7) til påvisning af bakterier, omfattende mindst to rum, der er adskilt fra hinanden med et filter (4), hvor et rum er et fagreservoir (8), der indeholder en suspension (1), der omfatter mindst en species af markerede testbakteriofager (2), der speci fikt binder til en bakterieart, der skal påvises, hvor markeringen er kendetegnende for de respektive arter af testbakteriofager (2), og et rum er et opsamlingsreservoir (9) til at optage filtratet efter tilsætning af en prøve (3) til suspensionen (1) og filtrering gennem filteret (4), og hvor filteret (4) har en porestørrelse på 0,1 pm til 0,5 pm, og porestørrelsen af det anvendte filter er valgt således, at den mindst ene bakterieart, der skal påvises, samt bakterie-bakteriofagkomplekser (5), der består af bakterier af den mindst ene bakterieart, der skal påvises, og dertil bundne testbakteriofager (2) af den mindst ene species, tilbageholdes og immo-biliseres på filteroverfladen, mens ubundne bakteriofager (2, 6) kan passere filteret; et middel (10) til identifikation af reaktionsbeholderen og mindst to målevinduer (11, 12), hvor et målevindue (11) er anbragt i området af den filteroverflade, der er vendt mod fagreservoiret (8), og et målevindue (12) er anbragt i området af opsamlingsreservoiret (9), kendetegnet ved, at suspensionen (1) endvidere omfatter en markeret reference-bakteriofag (6), der ikke binder til nogen bakterieart, der skal påvises, og en af testbakteriofagerne (2) udviser en disjunkt markering, eller at reaktionsbeholderen (7) omfatter et yderligere rum mellem filter (4) og fagreservoir (8), der indeholder en indikatoropløsning (28) med markerede mikro- eller nanopartikler, der ikke binder til nogen bakterieart, der skal påvises, og på grund af deres lille størrelse kan passere filteret (4).
- 10. Reaktionsbeholder (7) ifølge krav 9, der endvidere omfatter et rum mellem fagreservoir (8) og filter (4), der indeholder mindst et forfilter, hvor der ved det mindst ene forfilter er tale om en gelmatrix (24).
- 11. Patron, omfattende to eller flere reaktionsbeholdere (7) ifølge et af kravene 9-10, der er anbragt parallelt med hinanden, hvor hver reaktionsbeholder (7) indeholder en forskellig species af markerede testbakteriofager (2) i suspensionen (1) i fagreservoiret (8), der hver især binder specifikt til forskellige bakteriearter, der skal påvises, hvor markeringen af species er disjunkt.
- 12. Anvendelse af en reaktionsbeholder (7) ifølge et af kravene 9 til 10, eller en patron ifølge krav 11 til udøvelse af fremgangsmåden til påvisning af bakterier ifølge et af kravene 1 til 8.
- 13. Anvendelse af et måleapparat (15) til udøvelse af fremgangsmåden til påvisning af bakterier ifølge et af kravene 1 til 8, hvor måleapparatet (15) omfatter: en skakt, hvori indskydes en reaktionsbeholder (7) ifølge et af kravene 9-10 eller en patron ifølge krav 11; mindst to detektionsoptikker (16), der hver især er anbragt i området af målevinduerne (11, 12) i den indskudte reaktionsbeholder (7) eller patronen, hvor detektionsoptikkerne (16) hver især omfatter et linsesystem (17) til fokusering af emitteret lys fra markeringen af bakteriofagerne (2, 6) på mindst en optisk sensor (18), hvor den mindst ene optiske sensor (18) detekterer det emitterede lys; en processor, der i hvert enkelt tilfælde er forbundet med den mindst ene optiske sensor (18) i detektionsoptikkerne (16), og som behandler de modtagne detektionssignaler; og en med processoren forbundet udlæsningsenhed, der udlæser detektions-resultaterne til en bruger.
- 14. Anvendelse af et måleapparat ifølge krav 13, hvor detektionsoptikkerne (18) i måleapparatet (15) endvidere hver især omfatter en belysningsenhed (19), der emitterer lys i bølgelængdeområdet på 200-1000 nm, og et linsesystem (20) til fokusering af det af belysningsenheden (19) emitterede lys på et måleområde, hvor belysningsenheden (19) ligeledes er forbundet med processoren og styres af denne.
- 15. Anvendelse af et måleapparat ifølge et af kravene 13 eller 14, hvor måleapparatet (15) endvidere omfatter et integreret læseapparat, der registrerer midlet (10) til identifikation af reaktionsbeholderen (7) og videregiver de registrerede data til processoren, hvor processoren behandler de registrerede data, og resultaterne af identifikationen udlæses til en bruger via udlæsningsenheden.
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WO2006105414A2 (en) * | 2005-03-31 | 2006-10-05 | Colorado School Of Mines | Apparatus and method for detecting microscopic organisms using microphage |
CN1970786A (zh) * | 2005-11-23 | 2007-05-30 | 阮信畅 | 一种快速检测细菌的方法及装置 |
BRPI0810516A2 (pt) * | 2007-04-24 | 2014-10-07 | Biomatrica Inc | Armazenamento de amostra para ciência da vida |
CN101464409B (zh) * | 2007-12-19 | 2013-10-30 | 中国科学院电子学研究所 | 快速定量检测细菌的装置及方法 |
US9194859B2 (en) * | 2011-12-23 | 2015-11-24 | Abbott Point Of Care Inc. | Reader devices for optical and electrochemical test devices |
EP3011062A4 (en) * | 2013-06-19 | 2017-06-14 | Sample6 Technologies Inc. | Phage-based bacterial detection assay |
EP2905607A1 (en) * | 2014-02-07 | 2015-08-12 | Nxp B.V. | Analyte detection methods and devices |
-
2015
- 2015-11-20 PT PT15003307T patent/PT3171172T/pt unknown
- 2015-11-20 ES ES15003307.4T patent/ES2694874T3/es active Active
- 2015-11-20 DK DK15003307.4T patent/DK3171172T3/da active
- 2015-11-20 EP EP15003307.4A patent/EP3171172B1/de not_active Not-in-force
-
2016
- 2016-11-18 EP EP16798628.0A patent/EP3377902B1/de active Active
- 2016-11-18 CN CN201680079141.4A patent/CN108474792B/zh not_active Expired - Fee Related
- 2016-11-18 US US15/777,662 patent/US10766030B2/en active Active
- 2016-11-18 WO PCT/EP2016/001925 patent/WO2017084754A1/de active Application Filing
- 2016-11-18 RU RU2018117559A patent/RU2704245C1/ru not_active IP Right Cessation
- 2016-11-18 JP JP2018545543A patent/JP6691610B2/ja not_active Expired - Fee Related
Also Published As
Publication number | Publication date |
---|---|
PT3171172T (pt) | 2018-11-09 |
ES2694874T3 (es) | 2018-12-27 |
EP3171172A1 (de) | 2017-05-24 |
WO2017084754A1 (de) | 2017-05-26 |
JP2018535700A (ja) | 2018-12-06 |
US20180339293A1 (en) | 2018-11-29 |
EP3377902B1 (de) | 2020-09-16 |
EP3171172B1 (de) | 2018-07-18 |
CN108474792A (zh) | 2018-08-31 |
RU2704245C1 (ru) | 2019-10-25 |
JP6691610B2 (ja) | 2020-04-28 |
EP3377902A1 (de) | 2018-09-26 |
CN108474792B (zh) | 2021-02-02 |
US10766030B2 (en) | 2020-09-08 |
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