WO2000075636B1 - Multispectral taxonomic identification - Google Patents
Multispectral taxonomic identificationInfo
- Publication number
- WO2000075636B1 WO2000075636B1 PCT/US2000/015384 US0015384W WO0075636B1 WO 2000075636 B1 WO2000075636 B1 WO 2000075636B1 US 0015384 W US0015384 W US 0015384W WO 0075636 B1 WO0075636 B1 WO 0075636B1
- Authority
- WO
- WIPO (PCT)
- Prior art keywords
- amended
- probe
- cell
- probes
- cells
- Prior art date
Links
Classifications
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/6428—Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes"
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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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- 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
- C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
- C12Q1/6888—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for detection or identification of organisms
- C12Q1/689—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for detection or identification of organisms for bacteria
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/17—Systems in which incident light is modified in accordance with the properties of the material investigated
- G01N21/25—Colour; Spectral properties, i.e. comparison of effect of material on the light at two or more different wavelengths or wavelength bands
- G01N21/27—Colour; Spectral properties, i.e. comparison of effect of material on the light at two or more different wavelengths or wavelength bands using photo-electric detection ; circuits for computing concentration
- G01N21/274—Calibration, base line adjustment, drift correction
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/645—Specially adapted constructive features of fluorimeters
- G01N21/6456—Spatial resolved fluorescence measurements; Imaging
- G01N21/6458—Fluorescence microscopy
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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
- C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
- C12Q1/6888—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for detection or identification of organisms
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N2021/6417—Spectrofluorimetric devices
- G01N2021/6419—Excitation at two or more wavelengths
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N2021/6417—Spectrofluorimetric devices
- G01N2021/6421—Measuring at two or more wavelengths
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/645—Specially adapted constructive features of fluorimeters
- G01N2021/6463—Optics
- G01N2021/6471—Special filters, filter wheel
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2201/00—Features of devices classified in G01N21/00
- G01N2201/12—Circuits of general importance; Signal processing
- G01N2201/127—Calibration; base line adjustment; drift compensation
Landscapes
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Physics & Mathematics (AREA)
- Analytical Chemistry (AREA)
- Organic Chemistry (AREA)
- Immunology (AREA)
- General Health & Medical Sciences (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Engineering & Computer Science (AREA)
- Biochemistry (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- General Physics & Mathematics (AREA)
- Pathology (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Microbiology (AREA)
- General Engineering & Computer Science (AREA)
- Biophysics (AREA)
- Genetics & Genomics (AREA)
- Molecular Biology (AREA)
- Biotechnology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Mathematical Physics (AREA)
- Theoretical Computer Science (AREA)
- Spectroscopy & Molecular Physics (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Optics & Photonics (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
- Investigating, Analyzing Materials By Fluorescence Or Luminescence (AREA)
Abstract
The present invention provides an instrument and methods for a multispectral optical technique that can simultaneously classify individual biological cells within mixed populations. This invention, known as Multispectral Taxonomic Identification (MTID), shows that microscopy can be combined with a software analysis program to measure and categorize the fluorescence and other spectroscopically identifiable signals from complex populations of cells in situ , without cultivation. The invention thus enables high-throughput screening of cells for taxonomic classification.
Claims
1. (Amended) A method for empirically calibrating an optical system, the method comprising the steps of: (a) collecting data for calibration in a matrix [G], columns 1 through n of the matrix representing spectral channels, and rows 1 through of the matrix representing spectral groups; (b) solving for a correction matrix [C] ; (c) collecting vector data [Y] for pixels in an image representing the uncorrected intensity of the pixels in each of the 1 through n spectral channels of matrix [G]; [and ] (d) correcting [Y] by matrix multiplication with [C] to obtain empirically calibrated vectors [X] for the pixels m the calibrated imager.l;.and e further correcting the vectors [X\ to yield vectors \N\ by normalizing the vectors X\ to signals in a specific spectral channel.
2. The method of claim 1, where m equals n in matrix [G], and the correction matrix [C] is the inverse of [G]- 3. The method of claim 1 , where m is not equal to n in matrix [G], and the correction matrix [C] is solved for by singular value decomposition. 4. (Canceled) The method of claim 1 , wherein the vectors [X] are further corrected to yield vectors [N] by normalizing the vectors [X] to signals in a specific spectral channel. [5. j, (Amended) The method of claim 1, wherein the optical system is a microsjcope. [6.] 5. (Amended) The method of claim [5] 4, wherein the microscope operates in an epifluoresceπt configuration. [7.] 6. (Amended) The method of claim l, wherein the spectral channels are determined by filter configurations within an epi fluorescence microscope.
[8.] 7. (Amended) The method of claim 1 , wherein the spectral groups correspond to a set of fluoxophores used to identify a set of probes. [9.] 8. (Amended) The method of claim 1, where the calibration datais obtiined from a calibration sample. [ 10.] 2i (Amended) The method of claim 1 , wherein the image is of a sample comprising cells. [11.] i (Amended) The method of claim [10 3]] 9, wherein the cells are merhbers of the same taxa. [12.] (Amended) The method of claim [10 )]] 9, wherein a plurality of prob1 es is bound to the cells. [1 .] J (Amended) The method of claim [ 12 l]] IX, wherein the probes comprise oligonucleotides. [14.] 13. (Amended) The method of claim [135]] 12, wherein the o lig^nucleotide are deoxyoli gonucleo tides . [15.] 14, (Amended) The method of claim [12 -]] 11, wherein the probes are label|ed with fluorescent molecules. 116.1 15. (Amended) The method of claim [12 I]] I V, wherein the probes comprise antibodies. [17.] 16. (Amended) The method of claim [135]] 12, wherein the probes hybrijdize to rRNA. [18.] 17, (Amended) The method of claim [177]] 16, wherein the probes hybridize to 16S rRNA. [19.] 18. (Amended) The method of claim [177]] 16, wherein the probes hybrijdize to 18S rRNA. [20.1 19. (Amended) The method of claim [177]] 16, wherein the probes hybridize to 23S rRNA. [21.] 20. (Amended) The method of claim [177]] 16, wherein the probes hybridize to 28S rRNA- [22.] 21 (Amended) A method for empirically correcting images of cells, said images generated through the use of multiple spectral tags, comprising the steps of:
(a) selecting a probe that binds to a target cell; (b) dividing a sample of target cells into aliquots of target cells; (c) dividing a sample of probe solution into aliquots of probe solution; (d) labeling each aliquot of probe solution with a different spectral tag to generate a set of labeled probe solutions, wherein each member of the set is labeled with a different spectral tag; (e) mixing an aliquot of each member of the set of labeled probe solutions with an aliquot of target cells to generate a set of probe-iabeled target cells; (f) mixing together the probe-labeled target cells in approximately equal proportions; (g) acquiring images of the mixed probe-labeled target cells at multiple wavelengths; (h) calculating a correction matrix for each labeled probe set; and (i) applying the correction matrix to images of unknown cells to correct the image ofthe unknown cells. [23.] 22. (Amended) A method for taxonomic identification of a cell, comprising the steps of: (a) hybridizing at least one probe to a cell and quantifying the resulting first hybridization signal; (b) hybridizing the probe to at least one benchmark cell and quantifying the resulting second hybridization signal; and (c) taxonomically identifying the cell by calculating the difference between the second and first hybridization signals, wherein the difference increases with increasing taxonomic distance between the cell and the benchmark cell. [24.] 23. (Amended) The method of claim [23] 22. wherein the benchmark cell is) a known cell. [25.] 24. (Amended) The method of claim [23] 22, wherein the melting temperature, Tm! , characteristic of the hybridization reaction between the probe and
the cell is less than the melting temperature, T-^, characteristic of the hybridization reabtion between the probe and the benchmark cell.
1 [26.] 25. (Amended) The method of claim [23] 22, wherein the probe hybridizes to RNA.
1 [27.] 26. (Amended) The method of claim [26] 25, wherein the RNA is
1 [28.1 27. (Amended) The method of claim [23] 22, wherein the number of hybjridization probes is greater than one.
1 [29.] 28. (Amended) The method of claim [28] 27, wherein the probes are
2 desijgned so that the melting temperature, ^ characteristic of the hybridization
3 reacjtion between each probe and its complement is approximately equal.
I [30.] 29, (Amended) The method of claim [28] T7, wherein the probes are i desijgned to bind to one or more chromosomes of the cell.
1 [31.] 30. (Amended) The method of claim [28] 27, wherein the hybridization
2 signals originating from the hybridization between the probes and the cell generate a
3 muli|i spectral fingerprint of the cell's taxon.
1 [32.] 3T (Amended) The method of claim [23] 22, wherein the quantifying
2 step |(b) resolves more than two levels of signal intensity.
1 [ 3. ] 32, (Amended) The method of claim [23 ] 22, wherein the quantifying
2 step (b) resolves more than four levels of signal intensity.
1 [34,] 33. (Amended) The method of claim [23] 22, wherein the quantifying
2 step (b) resolves more than eight levels of signal intensity.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| AU55952/00A AU5595200A (en) | 1999-06-04 | 2000-06-02 | Multispectral taxonomic identification |
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| US13745899P | 1999-06-04 | 1999-06-04 | |
| US60/137,458 | 1999-06-04 |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| WO2000075636A1 WO2000075636A1 (en) | 2000-12-14 |
| WO2000075636B1 true WO2000075636B1 (en) | 2001-02-08 |
Family
ID=22477524
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| PCT/US2000/015384 WO2000075636A1 (en) | 1999-06-04 | 2000-06-02 | Multispectral taxonomic identification |
Country Status (2)
| Country | Link |
|---|---|
| AU (1) | AU5595200A (en) |
| WO (1) | WO2000075636A1 (en) |
Families Citing this family (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20040243318A1 (en) * | 2001-07-18 | 2004-12-02 | Akio Ogawa | Microbe examining device and method |
| EP1523580A2 (en) * | 2002-07-18 | 2005-04-20 | Henkel Kommanditgesellschaft auf Aktien | Detection of microorganisms |
| BR0312944A (en) * | 2002-07-18 | 2005-07-12 | Henkel Kommanditgesellchaft Au | Oligonucleotides for microorganism detection |
| ES2368321B2 (en) * | 2011-03-31 | 2013-06-07 | Universidad Politécnica de Madrid | METHOD FOR OBTAINING MULTIESPECTRAL IMAGES OF ABSOLUTE REFLECTANCE. |
| BE1024355B1 (en) * | 2016-06-30 | 2018-02-05 | Container Quick Lock Luxembourg Sa | DETERMINATION OF MICROBIAL COMMUNITIES AND BIOFILMS |
| LU93141B1 (en) * | 2016-06-30 | 2018-01-09 | Container Quick Lock Luxembourg S A | Determination of microbial communities and biofilms |
| WO2018175377A1 (en) | 2017-03-24 | 2018-09-27 | Axon Dx, Llc | Spectral imaging apparatus and methods |
| IT201800004146A1 (en) * | 2018-03-30 | 2019-09-30 | St Microelectronics Srl | APPARATUS FOR ANALYSIS OF MOLECULES BY DETECTION OF FLUORESCENT SIGNALS AND RELATIVE METHOD |
| US20220236187A1 (en) * | 2019-06-07 | 2022-07-28 | Helmholtz Zentrum München - Deutsches Forschungszentrum für Gesundheit und Umwelt (GmbH) | System and method for real-time multicolor shortwave infrared fluorescence imaging |
| CN117237352B (en) * | 2023-11-14 | 2024-02-09 | 之江实验室 | Gene chip image processing method, data processing device and storage medium |
Family Cites Families (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5723597A (en) * | 1983-01-10 | 1998-03-03 | Gen-Probe Incorporated | Ribosomal nucleic acid probes for detecting organisms or groups of organisms |
| US4660151A (en) * | 1983-09-19 | 1987-04-21 | Beckman Instruments, Inc. | Multicomponent quantitative analytical method and apparatus |
| US6015667A (en) * | 1996-06-03 | 2000-01-18 | The Perkin-Emer Corporation | Multicomponent analysis method including the determination of a statistical confidence interval |
| CA2291730A1 (en) * | 1997-06-05 | 1998-12-10 | Kairos Scientific Inc. | Calibration of fluorescence resonance energy transfer in microscopy |
-
2000
- 2000-06-02 WO PCT/US2000/015384 patent/WO2000075636A1/en active Application Filing
- 2000-06-02 AU AU55952/00A patent/AU5595200A/en not_active Abandoned
Also Published As
| Publication number | Publication date |
|---|---|
| WO2000075636A1 (en) | 2000-12-14 |
| AU5595200A (en) | 2000-12-28 |
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