WO2000015832A1 - Detection of cells in a liquid sample - Google Patents
Detection of cells in a liquid sample Download PDFInfo
- Publication number
- WO2000015832A1 WO2000015832A1 PCT/DE1999/002867 DE9902867W WO0015832A1 WO 2000015832 A1 WO2000015832 A1 WO 2000015832A1 DE 9902867 W DE9902867 W DE 9902867W WO 0015832 A1 WO0015832 A1 WO 0015832A1
- Authority
- WO
- WIPO (PCT)
- Prior art keywords
- cells
- sample
- added
- filtered
- detection
- Prior art date
Links
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
- C12Q1/02—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving viable microorganisms
- C12Q1/04—Determining presence or kind of microorganism; Use of selective media for testing antibiotics or bacteriocides; Compositions containing a chemical indicator therefor
- C12Q1/06—Quantitative determination
Definitions
- the invention relates to a method for the detection of cells in a sample in liquid form, in which a substance is added to the sample which leads to the formation of a specific enzyme in active cells, and the cells are filtered off with a membrane filter.
- Such a method is known from DE 38 55 762 Tl.
- the coliform bacteria present in a liquid are concentrated using a filter, the pores of which are sufficiently small to retain the bacteria.
- This filter is then placed on a nutrient medium to increase the number of cells.
- This proliferation is promoted in such a way that a microcolony visible to each individual bacterium is formed under fluorescence conditions.
- These microcolonies are then irradiated with light of a certain wavelength so that the number of fluorescent microcolonies can be counted.
- the invention is therefore based on the object of providing a method in which a much faster detection of cells capable of reproduction in a sample is possible.
- This object is achieved with a generic method in which, immediately after filtering off the cells with a membrane filter, the filtered cells are examined fluorescence-optically with the aid of a microscope.
- the advantage of the method is that this examination can be carried out on a single cell.
- the sample drawn does not therefore have to be enriched or multiplied, as in classic microbiology, in order to subsequently examine a cell population.
- the sample is examined directly after the membrane filtration without an intermediate growth step, and the method allows information about the enzyme formation capacity of a cell in the sample after only one hour.
- the fluorescent dye can be added at any stage of the process before the microscopic examination. In various applications, however, it is advantageous if the fluorescent dye follows the filtration is added to the cells filtered off since this can reduce the background radiation.
- a polycarbonate filter is advantageously used to separate the cells from the sample. These filters are particularly well suited to filtering off the microorganism sought by choosing an appropriate pore size.
- An advantageous embodiment provides that a substance is added to the cells, which opens the cell walls at least in places. This makes it easier for the inducer of the enzyme sought and the fluorescent dye to enter the cell.
- the method according to the invention is particularly suitable for food samples. It is advantageous if the sample is a liquid or liquefied food.
- the method is particularly suitable for the analysis of beverages, such as preferably beer or non-alcoholic beverages.
- the first process step is to convert the sample material to be examined into a liquid, aqueous form. If foods such as cheese or jam are to be examined, these substances must be liquefied in a suitable manner. In the figure the substance to be examined is designated with the reference number 1.
- the aqueous form of the sample material to be examined in the Erlenmeier flask has the reference number 2. If the sample is a drink, the addition of the sample material is unnecessary since the sample 2 is already in liquid form.
- the inducer 3 of the enzyme sought is added to this sample.
- an inducer for galactosidase e.g. Galactose added.
- a substance 5 can also be added subsequently or simultaneously, which opens the cell walls of the sought-after microorganisms at different locations.
- Mutanolysin for example, can be used as the cell wall-opening substance.
- cell wall-accessible substances that can pass through the cell wall without digestion.
- the sample After an exposure time and exposure temperature corresponding to the substrate, the sample is filtered through a polycarbonate membrane filter 6 with a pore size corresponding to the microorganism sought. The sample thereby arrives in the collecting container 7 and the microorganisms sought remain on the membrane filter 8.
- a fluorescent dye 4 is added to the filtered cells remaining on the filter.
- a reagent specific for galactosidase such as fluorescindigalactoside, is added.
- the membrane filter 8 is then examined fluorescence optically. This is carried out with the aid of a microscope 9. In microscopic image 10, the cells which have the enzyme activity sought are illuminated according to the fluorescent dye.
- the number of luminous dots or the intensity of the luminosity can be used to easily deduce the contamination of the sample with lactobacilli capable of reproduction.
- the result takes into account both the number of microorganisms per sample volume and the enzyme activity and thus the ability of the cells found to proliferate.
Landscapes
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Organic Chemistry (AREA)
- Engineering & Computer Science (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Analytical Chemistry (AREA)
- Toxicology (AREA)
- Immunology (AREA)
- Microbiology (AREA)
- Molecular Biology (AREA)
- Biophysics (AREA)
- Physics & Mathematics (AREA)
- Biotechnology (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
Description
Claims
Priority Applications (3)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
DE19981818T DE19981818D2 (en) | 1998-09-11 | 1999-09-09 | Detection of cells in a liquid sample |
AU11493/00A AU1149300A (en) | 1998-09-11 | 1999-09-09 | Detection of cells in a liquid sample |
EP99969120A EP1119638A1 (en) | 1998-09-11 | 1999-09-09 | Detection of cells in a liquid sample |
Applications Claiming Priority (2)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
DE19841588.5 | 1998-09-11 | ||
DE1998141588 DE19841588A1 (en) | 1998-09-11 | 1998-09-11 | Detecting cells especially in food or drink by adding a material giving rise to an enzyme followed by optical fluorescence examination |
Publications (1)
Publication Number | Publication Date |
---|---|
WO2000015832A1 true WO2000015832A1 (en) | 2000-03-23 |
Family
ID=7880617
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
PCT/DE1999/002867 WO2000015832A1 (en) | 1998-09-11 | 1999-09-09 | Detection of cells in a liquid sample |
Country Status (4)
Country | Link |
---|---|
EP (1) | EP1119638A1 (en) |
AU (1) | AU1149300A (en) |
DE (2) | DE19841588A1 (en) |
WO (1) | WO2000015832A1 (en) |
Families Citing this family (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
DE102004011822A1 (en) * | 2004-03-11 | 2005-09-29 | Cognis Deutschland Gmbh & Co. Kg | Quality assurance system for the detection of microorganisms |
Citations (6)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
EP0101398A1 (en) * | 1982-07-21 | 1984-02-22 | Packard Instrument Company, Inc. | Method of concentrating and measuring unicellular organisms |
WO1989004372A1 (en) * | 1987-11-05 | 1989-05-18 | Berg James D | Rapid process for detecting pathogenic microorganisms |
US5510243A (en) * | 1994-06-21 | 1996-04-23 | Gelman Sciences, Inc. | Multiple chromogen enzyme targeting (MCET) for use in bacterial contamination monitoring |
WO1996014431A1 (en) * | 1994-11-07 | 1996-05-17 | Studie-En Samenwerkingverband Vlaams Water | Enzymatic method for detecting coliform bacteria or e. coli |
DE19608320A1 (en) * | 1996-02-22 | 1997-08-28 | Biosquant Gmbh | Rapid, high- sensitivity determination of microbial quality of water |
WO1997049830A1 (en) * | 1996-06-27 | 1997-12-31 | Isrim S.C.A.R.L. | Method for detecting cells in a sample using an enzyme induction promoter |
Family Cites Families (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US4242447A (en) * | 1978-11-29 | 1980-12-30 | Bioresearch | Rapid detection of bacteria |
-
1998
- 1998-09-11 DE DE1998141588 patent/DE19841588A1/en not_active Withdrawn
-
1999
- 1999-09-09 DE DE19981818T patent/DE19981818D2/en not_active Expired - Fee Related
- 1999-09-09 WO PCT/DE1999/002867 patent/WO2000015832A1/en not_active Application Discontinuation
- 1999-09-09 AU AU11493/00A patent/AU1149300A/en not_active Abandoned
- 1999-09-09 EP EP99969120A patent/EP1119638A1/en not_active Withdrawn
Patent Citations (7)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
EP0101398A1 (en) * | 1982-07-21 | 1984-02-22 | Packard Instrument Company, Inc. | Method of concentrating and measuring unicellular organisms |
WO1989004372A1 (en) * | 1987-11-05 | 1989-05-18 | Berg James D | Rapid process for detecting pathogenic microorganisms |
DE3852825T2 (en) * | 1987-11-05 | 1995-05-18 | James D Berg | QUICK PROCEDURE FOR DETECTING COLIFORM MICROORGANISMS. |
US5510243A (en) * | 1994-06-21 | 1996-04-23 | Gelman Sciences, Inc. | Multiple chromogen enzyme targeting (MCET) for use in bacterial contamination monitoring |
WO1996014431A1 (en) * | 1994-11-07 | 1996-05-17 | Studie-En Samenwerkingverband Vlaams Water | Enzymatic method for detecting coliform bacteria or e. coli |
DE19608320A1 (en) * | 1996-02-22 | 1997-08-28 | Biosquant Gmbh | Rapid, high- sensitivity determination of microbial quality of water |
WO1997049830A1 (en) * | 1996-06-27 | 1997-12-31 | Isrim S.C.A.R.L. | Method for detecting cells in a sample using an enzyme induction promoter |
Also Published As
Publication number | Publication date |
---|---|
AU1149300A (en) | 2000-04-03 |
DE19841588A1 (en) | 2000-03-23 |
DE19981818D2 (en) | 2001-09-27 |
EP1119638A1 (en) | 2001-08-01 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
DE3855762T2 (en) | Direct method to detect very low levels of coliform contamination | |
DE69322073T2 (en) | ANALYSIS PROCEDURE | |
DE2823916C2 (en) | ||
DE69020555T2 (en) | Precipitation test for microorganisms. | |
DE69419948T2 (en) | Method for determining the number of living microorganisms using a hydrophobic membrane filter | |
DE69434255T2 (en) | NUCLEIC ACID PROBE FOR THE DETECTION OF LACTOBACILLUS | |
DE69619395T2 (en) | MULTI-ZONE STERILITY INDICATOR | |
EP1725676B2 (en) | Measuring contamination | |
DE69512037T2 (en) | METHOD FOR CONDITIONING LIQUID SAMPLES | |
DE102009033368A1 (en) | Mass spectrometric sepsis diagnosis | |
EP1846569B1 (en) | Method for identifying germs | |
DE69720248T2 (en) | FAST MICROBIAL TEST | |
DE3903777A1 (en) | METHOD FOR THE QUICK DETECTION OF MICROORGANISMS IN SAMPLES AND DEVICE FOR IMPLEMENTING THE METHOD | |
DE69610537T2 (en) | Methods for the detection of microorganisms | |
DE60035977T2 (en) | FTA-COATED CARRIER FOR USE AS A MOLECULAR DIAGNOSTIC | |
EP1066403A1 (en) | Method for detecting micro-organisms in gases | |
WO2004055203A1 (en) | Method and device for identifying germs | |
DE68909445T2 (en) | Method for determining genetic sequences and test set therefor. | |
DE60130970T2 (en) | REAGENT SET FOR DETECTING MICROORGANISMS, APPARATUS FOR QUANTIFYING MICROORGANISMS AND METHOD FOR QUANTIFYING MICROORGANISMS | |
EP3872186A1 (en) | Method for spectrometric characterization of microorganisms | |
WO2000015832A1 (en) | Detection of cells in a liquid sample | |
DE69214473T2 (en) | A method for the detection of gram-negative bacteria | |
EP0342501A2 (en) | Method and means for the determination of bacteria or somatic cells in milk | |
DE69214474T2 (en) | A method for the detection of microorganisms | |
DE3784702T2 (en) | METHOD AND DEVICE FOR DETECTING AND QUANTIFYING BACTERIA. |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
ENP | Entry into the national phase |
Ref document number: 2000 11493 Country of ref document: AU Kind code of ref document: A |
|
AK | Designated states |
Kind code of ref document: A1 Designated state(s): AE AL AM AT AU AZ BA BB BG BR BY CA CH CN CR CU CZ DE DK DM EE ES FI GB GD GE GH GM HR HU ID IL IN IS JP KE KG KP KR KZ LC LK LR LS LT LU LV MD MG MK MN MW MX NO NZ PL PT RO RU SD SE SG SI SK SL TJ TM TR TT UA UG US UZ VN YU ZA ZW |
|
AL | Designated countries for regional patents |
Kind code of ref document: A1 Designated state(s): GH GM KE LS MW SD SL SZ UG ZW AM AZ BY KG KZ MD RU TJ TM AT BE CH CY DE DK ES FI FR GB GR IE IT LU MC NL PT SE BF BJ CF CG CI CM GA GN GW ML MR NE SN TD TG |
|
DFPE | Request for preliminary examination filed prior to expiration of 19th month from priority date (pct application filed before 20040101) | ||
121 | Ep: the epo has been informed by wipo that ep was designated in this application | ||
WWE | Wipo information: entry into national phase |
Ref document number: 1999969120 Country of ref document: EP |
|
WWP | Wipo information: published in national office |
Ref document number: 1999969120 Country of ref document: EP |
|
REF | Corresponds to |
Ref document number: 19981818 Country of ref document: DE Date of ref document: 20010927 |
|
WWE | Wipo information: entry into national phase |
Ref document number: 19981818 Country of ref document: DE |
|
WWW | Wipo information: withdrawn in national office |
Ref document number: 1999969120 Country of ref document: EP |