WO1996007759A1 - Reactifs de dosage et de bioluminescence - Google Patents

Reactifs de dosage et de bioluminescence Download PDF

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Publication number
WO1996007759A1
WO1996007759A1 PCT/GB1995/002085 GB9502085W WO9607759A1 WO 1996007759 A1 WO1996007759 A1 WO 1996007759A1 GB 9502085 W GB9502085 W GB 9502085W WO 9607759 A1 WO9607759 A1 WO 9607759A1
Authority
WO
WIPO (PCT)
Prior art keywords
medium
acid
luciferase
kit according
luciferin
Prior art date
Application number
PCT/GB1995/002085
Other languages
English (en)
Inventor
Nicholas Peter Martin Foote
Michael Noble
Peter Leonard Grant
Christopher Thomas Evans
Original Assignee
Celsis International Plc
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 Celsis International Plc filed Critical Celsis International Plc
Priority to AU33949/95A priority Critical patent/AU3394995A/en
Publication of WO1996007759A1 publication Critical patent/WO1996007759A1/fr

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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
    • C12Q1/66—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving luciferase

Definitions

  • BIOLUMINESCENCE AND ASSAY REAGENTS This invention relates to a method for detecting microorganisms.
  • Live microorganisms can be detected with great « sensitivity by measuring their ATP using a reagent based on the firefly luciferase reaction. This ATP must be extracted by lysing the microorganisms. There are presently four main ways of doing this, i.e. extracting with boiling buffer or with detergent, or using solvents or acids.
  • the detergent method is understandably the most popular because it involves a minimum of sample handling, is almost instantaneous, and allows a simple two-step assay, comprising: (1) adding an extractant to the sample; and (2) adding reagent and measuring light.
  • This method is the subject of GB-A-1604249.
  • the detergent often actually stimulates the initial light production, but also rapidly inactivates the enzyme. Thus the light output falls to nothing over a period of a few minutes. Therefore, despite the initial stimulation, there may be a drastic loss of assay sensitivity, especially if the light output is not measured straight away.
  • non-ionic detergents protect, at least in part, from this inactivation. However, they may also protect bacteria from the lytic effects of the cationic detergent; therefore, a ⁇ 'mixed" detergent extractant is not efficient. Furthermore, non-ionic detergents themselves have a slow inactivating effect on luciferase and cannot be included as part of a stable reagent.
  • An object behind the present invention is to provide a reagent having a low decay rate, i.e. ⁇ 5% per minute. e.g. even after the addition of a detergent-based extractant, so that it can be used in a two-step assay.
  • This could be used in even the simplest luminometer, e.g. a camera-type microtitre plate system, and would bring all 'the advantages of improved accuracy and sensitivity of a true low-decay rate assay.
  • medium chain- length fatty acids e.g. C 5 . 15 alkanoic acids
  • Their esters or other derivatives/analogues may also be used.
  • a reagent containing octanoic acid retains low-decay rate kinetics even in the presence of an efficient extractant based on cationic detergents. It has little effect on the storage stability of luciferase and can even be added to a luciferase-luciferin reagent before it is freeze-dried, although the, say, octanoic acid is preferably included in the reagent reconstitution buffer. Such a reagent could still be used after 8 hours at 25 ⁇ C or 1 week at 4°C.
  • Example 1 illustrate the invention.
  • the latter comprised 50 mM Tris-Hepes, 9 mM magnesium acetate, 1 mM EDTA, 1 g/1 octanoic acid and 0.02% sodium azide. After incubation at 15°C for 1 hour, the two preparations were compared in an assay of ATP in the presence of a bacterial lysing reagent containing cationic detergents.
  • the assay was performed as follows using a Berthold Biolumat LB9500T luminometer: 0.1 ml of the lysing agent was placed in a cuvette, 0.1 ml of 2 x 10 *9 M Na j ATP in sterile water was added, and the cuvette was equilibrated at 25°C for 5 minutes. 0.1 ml of the reagent was then added. The initial light output was measured by a 10 second integration, and the light signal was continuously monitored on a chart recorded so that the decay rate could be calculated. The stability of the reconstituted reagents was compared by storage at 25°C and 4°C The results were as follows:

Landscapes

  • 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)

Abstract

Kit constitué par deux ou plusieurs récipients et servant à effectuer une évaluation de l'ATP libéré par des micro-organismes. Ce kit contient de la luciférine, de la luciférase, un milieu de reconstitution et, éventuellement, un tensioactif ou un autre agent de lyse cellulaire, ainsi que, de plus, un acide alcanoïque à chaîne moyenne.
PCT/GB1995/002085 1994-09-05 1995-09-04 Reactifs de dosage et de bioluminescence WO1996007759A1 (fr)

Priority Applications (1)

Application Number Priority Date Filing Date Title
AU33949/95A AU3394995A (en) 1994-09-05 1995-09-04 Bioluminescence and assay reagents

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
GB9417793.8 1994-09-05
GB9417793A GB9417793D0 (en) 1994-09-05 1994-09-05 Bioluminescence and assay reagents

Publications (1)

Publication Number Publication Date
WO1996007759A1 true WO1996007759A1 (fr) 1996-03-14

Family

ID=10760816

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/GB1995/002085 WO1996007759A1 (fr) 1994-09-05 1995-09-04 Reactifs de dosage et de bioluminescence

Country Status (3)

Country Link
AU (1) AU3394995A (fr)
GB (1) GB9417793D0 (fr)
WO (1) WO1996007759A1 (fr)

Cited By (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0816512A1 (fr) * 1996-06-24 1998-01-07 Basf Aktiengesellschaft Utilisation d'un essai de bioluminescence pour la détermination de microorganismes en dispersions contenant des polymères et/ou des pigments
EP1041151A1 (fr) * 1997-12-26 2000-10-04 Kikkoman Corporation Luciferase et technique permettant de doser l'atp intracellulaire au moyen de ladite luciferase
EP1048739A1 (fr) * 1997-12-03 2000-11-02 Kikkoman Corporation Methode d'analyse de composants intracellulaires
DE102011077238A1 (de) 2011-06-09 2012-12-13 Bayerische Motoren Werke Aktiengesellschaft Verfahren zum Nachweis von Mikroorganismen in einem Hohlraumkonservierungsmittel
US8609330B2 (en) 2008-12-31 2013-12-17 3M Innovative Properties Company Live Bioload detection using microparticles
US9284593B2 (en) 2009-12-30 2016-03-15 3M Innovative Properties Company Live bioload detection using microparticles

Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US3423290A (en) * 1966-08-03 1969-01-21 Nasa Lyophilized reaction mixtures
US3666631A (en) * 1969-12-31 1972-05-30 Nasa Bacterial contamination monitor
EP0309429A2 (fr) * 1987-09-23 1989-03-29 Life Science International Ab Détermination d'ATP cellulaire par luminescence
FR2699930A1 (fr) * 1992-12-24 1994-07-01 Kabore Paul Réactifs et procédés pour le dosage d'éléments réactifs et autres cofacteurs d'une réaction.

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US3423290A (en) * 1966-08-03 1969-01-21 Nasa Lyophilized reaction mixtures
US3666631A (en) * 1969-12-31 1972-05-30 Nasa Bacterial contamination monitor
EP0309429A2 (fr) * 1987-09-23 1989-03-29 Life Science International Ab Détermination d'ATP cellulaire par luminescence
FR2699930A1 (fr) * 1992-12-24 1994-07-01 Kabore Paul Réactifs et procédés pour le dosage d'éléments réactifs et autres cofacteurs d'une réaction.

Non-Patent Citations (3)

* Cited by examiner, † Cited by third party
Title
CHEMICAL ABSTRACTS, vol. 110, no. 15, 10 April 1989, Columbus, Ohio, US; abstract no. 131899, D. M. BYERS.: "Elongation of exogenous fatty acids by the bioluminescent bacterium Vibrio harveyi." page 402; column 1; *
J. BACTERIOL., vol. 171, no. 1, pages 59 - 64 *
W. A. FRANCISCO ET AL.: "Interaction of bacterial luciferase with aldehyde substrates and inhibitors.", JOURNAL OF BIOLOGICAL CHEMISTRY, vol. 268, no. 33, 25 November 1993 (1993-11-25), MD US, pages 24734 - 24741 *

Cited By (12)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0816512A1 (fr) * 1996-06-24 1998-01-07 Basf Aktiengesellschaft Utilisation d'un essai de bioluminescence pour la détermination de microorganismes en dispersions contenant des polymères et/ou des pigments
EP1048739A1 (fr) * 1997-12-03 2000-11-02 Kikkoman Corporation Methode d'analyse de composants intracellulaires
EP1048739A4 (fr) * 1997-12-03 2004-06-30 Kikkoman Corp Methode d'analyse de composants intracellulaires
EP1041151A1 (fr) * 1997-12-26 2000-10-04 Kikkoman Corporation Luciferase et technique permettant de doser l'atp intracellulaire au moyen de ladite luciferase
EP1041151A4 (fr) * 1997-12-26 2002-09-18 Kikkoman Corp Luciferase et technique permettant de doser l'atp intracellulaire au moyen de ladite luciferase
US6812012B1 (en) 1997-12-26 2004-11-02 Kikkoman Corporation Luciferase and methods for assaying intracellular ATP by using the same
US7560245B2 (en) 1997-12-26 2009-07-14 Kikkoman Corporation Luciferase and methods for measuring intracellular ATP using the same
US8609330B2 (en) 2008-12-31 2013-12-17 3M Innovative Properties Company Live Bioload detection using microparticles
US9382570B2 (en) 2008-12-31 2016-07-05 3M Innovative Properties Company Live bioload detection using microparticles
US9284593B2 (en) 2009-12-30 2016-03-15 3M Innovative Properties Company Live bioload detection using microparticles
DE102011077238A1 (de) 2011-06-09 2012-12-13 Bayerische Motoren Werke Aktiengesellschaft Verfahren zum Nachweis von Mikroorganismen in einem Hohlraumkonservierungsmittel
DE102011077238B4 (de) * 2011-06-09 2016-03-31 Bayerische Motoren Werke Aktiengesellschaft Verfahren zum Nachweis von Mikroorganismen in einem Hohlraumkonservierungsmittel

Also Published As

Publication number Publication date
GB9417793D0 (en) 1994-10-26
AU3394995A (en) 1996-03-27

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