EP1141724A1 - Nmr spectroscopy method - Google Patents

Nmr spectroscopy method

Info

Publication number
EP1141724A1
EP1141724A1 EP99963638A EP99963638A EP1141724A1 EP 1141724 A1 EP1141724 A1 EP 1141724A1 EP 99963638 A EP99963638 A EP 99963638A EP 99963638 A EP99963638 A EP 99963638A EP 1141724 A1 EP1141724 A1 EP 1141724A1
Authority
EP
European Patent Office
Prior art keywords
xenon
biological molecule
assay
nmr
hyperpolarised
Prior art date
Legal status (The legal status 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 status listed.)
Withdrawn
Application number
EP99963638A
Other languages
German (de)
French (fr)
Inventor
Peter Knox
Neil Cook
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
GE Healthcare Ltd
Original Assignee
Nycomed Amersham 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 Nycomed Amersham PLC filed Critical Nycomed Amersham PLC
Publication of EP1141724A1 publication Critical patent/EP1141724A1/en
Withdrawn legal-status Critical Current

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
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6813—Hybridisation assays
    • C12Q1/6816—Hybridisation assays characterised by the detection means
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/58—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving labelled substances
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01R—MEASURING ELECTRIC VARIABLES; MEASURING MAGNETIC VARIABLES
    • G01R33/00—Arrangements or instruments for measuring magnetic variables
    • G01R33/20—Arrangements or instruments for measuring magnetic variables involving magnetic resonance
    • G01R33/44—Arrangements or instruments for measuring magnetic variables involving magnetic resonance using nuclear magnetic resonance [NMR]
    • G01R33/46—NMR spectroscopy
    • G01R33/465—NMR spectroscopy applied to biological material, e.g. in vitro testing

Definitions

  • This invention is concerned with nuclear magnetic resonance (NMR) spectroscopy and/or 5 NMR imaging.
  • the technique involves observing the spectrum/image of a NMR active nuclear species in vitro in order to obtain information about the environment in which the species is present.
  • the spectra of NMR active nuclei vary depending on their environment, and this is reported in the literature (PNAS, 93,12932-6, 1996).
  • Noble gases having non-zero nuclear spin can be hyperpolarised, i.e. have their polarisation enhanced over the equilibrium polarisation, e.g. by the use of circularly polarised light.
  • Preferred techniques for hyperpolarisation include spin exchange with an optically pumped alkali metal vapour and metastability exchange.
  • Noble gases to which this technique can be applied include 3 He and 129 Xe. As described by M S Albert et al in US Patent 5,545,396, the
  • 15 technique can be used to prepare hyperpolarised noble gases that can be administered by inhalation for magnetic resonance imaging of the human body.
  • Xenon is chemically inert and has hydrophobic properties, and is capable of being weakly bound by hydrophobic regions of biological molecules (PNAS, 78, No 8, 4946-9, August 20 1981; Abstracts of the 11 ⁇ Annual Meeting of the Society for Magnetic Resonance in Medicine (1992) page 2104). Thus it is possible to "label" biological molecules with xenon.
  • This invention concerns the method of labelling biological molecules with hyperpolarised 129 Xe. All macromolecules have a number of discrete hydrophobic and hydrophilic sites. Xenon 25 will bind by hydrophobic interactions to hydrophobic sites with different affinity. The xenon labels the biological compound by becoming weakly bound to it, e.g. at specific hydrophobic sites on a surface of or within a cavity of a protein or other macromolecule.
  • the NMR sensitivity of hyperpolarised xenon is highly increased compared to non- 30 hyperpolarised xenon.
  • Another advantage of the present invention is the reversible and nondestructive nature of the bond between the xenon and the biological molecule.
  • a further advantage is that the forming of the "bond" and subsequent measurement may be repeated if needed.
  • xenon is a gas (condensation temperature of -106°C), it and may easily and rapidly be separated from the biological molecule if necessary.
  • xenon is essential chemically inert and will not adversely effect the biological molecule.
  • One embodiment of the invention thus provides an in vitro method which comprises labelling a biological molecule with hyperpolarised xenon, and observing a magnetic resonance spectrum and/or image of the hyperpolarised xenon in the environment of the biological molecule.
  • the spectrum/image provides information about the environment(s) at which atoms of xenon are bound to the biological molecule.
  • Any conformational change of the biological molecule resulting e.g. from the binding (or the disappearance) of a ligand (e.g. a lipid, carbohydrate, peptide, polypeptide, nucleic acid or any sort of drug) or cleavage by an enzyme will cause an alteration in the xenon NMR spectrum.
  • a ligand e.g. a lipid, carbohydrate, peptide, polypeptide, nucleic acid or any sort of drug
  • Each hydrophobic site in the biological molecule may give rise to a specific and characteristic NMR shift.
  • a further embodiment of the present invention is to take NMR "fmgerprint(s)" of a known biological molecule. These fingerprints can subsequently be used to identify unknowns by direct comparison in a manner similar to infra-red spectroscopy.
  • a biological molecule as defined by the present invention is a monomeric or polymeric molecule that is present in biological systems or that is artificially introduced and is biologically active in such systems.
  • Biological molecules include lipids, sugars and polysaccharides, nucleic acids (DNA, RNA), nucleosides, oligonucleosides, polynucleosides, nucleotides, oligonucleotides, polynucleotides, enzymes, vitamins and particularly peptides, polypeptides and proteins.
  • the labelled biological molecule is an assay reagent taking part in an assay method and wherein the assay reagent is labelled with hyperpolarised xenon.
  • the labelling of the biological molecule with hyperpolarised xenon may be performed before, during or after performance of the assay. ___.
  • An assay method is a test involving a reaction of one or more biological molecules.
  • the assays include for example competition assays (e.g. receptor- ligand antagonism, enzyme-substrate inhibitors, protein-protein interaction inhibitors), binding assays (e.g. receptor-ligand agonism, enzyme-substrate reactions, protein-protein interactions), immunoassays (e.g. for specific analytes), hybridisation assays (e.g. nuclease assays, mutation analysis, mRNA and DNA detection), test involving cells, organs and/or whole organisms. These tests may involve e.g.
  • Assays include binding studies performed on eukaryotic and prokaryotic microorganisms; binding studies performed on tissue in vitro; and binding studies in which an assay reagent is administered in vivo and an excretion product (e.g. urine, faeces, or breath) analysed by NMR in vitro.
  • an excretion product e.g. urine, faeces, or breath
  • Assays performed in vitro may conveniently be in multiwell plates, with either an assay reagent in the wells of the plate being labelled with hyperpolarised xenon, or a reagent being so labelled in bulk prior to being dispensed into individual wells of the plate.
  • the biological molecule is present in a liquid medium into which the xenon is introduced as a gas. This may be achieved e.g. by bubbling it through the fluid or by contact with the biological molecule as a solid.
  • the xenon is introduced as a solution in a solvent, which is compatible with the biological molecule (e.g. in a lipophilic solvent such as a lipid or a fluorocarbon solvent).
  • the liquid medium used according to one embodiment of the present invention may be deuterated water, deuterated buffers or solvents, e.g. lipophilic solvents which may contain lipid bicelles, lipid vesicles, liposomes, cryptophanes and/or cyclodextrins.
  • deuterated buffers or solvents e.g. lipophilic solvents which may contain lipid bicelles, lipid vesicles, liposomes, cryptophanes and/or cyclodextrins.
  • 129 Xe has a natural abundance of 26.4%.
  • the xenon used for this invention may be either the naturally occurring material or one artificially enriched in 12 Xe.
  • a preferred degree of enrichment 129 Xe is 40 % or more.
  • a more preferred degree is 50 % or more and an even more preferred degree is 75 % or more.
  • a particularly preferred degree of enrichment is 90 % or more.
  • Bulk supplies of xenon enriched in 129 Xe and hyperpolarised to a high degree are now available commercially and have a half life long enough to permit transport over substantial distances. While the half life of hyperpolarised I29 Xe in the biological environments contemplated in this invention will be lower, it is expected to be amply sufficient to permit the desired spectra to be obtained.
  • a preferred degree of hype ⁇ olarisation is 8 % or more.
  • a more preferred hype ⁇ olarisation degree is 20 % or more and an even more preferred degree is 30 % or more. Ideally, the degree will approach 100
  • the temperature at the time xenon is added is above the temperature at which the biological molecule is frozen, but below the temperature at which the biological molecule may be denatured.
  • xenon may be added to the frozen biological molecule, followed by thawing.
  • the right temperature to achieve the optimal function of the biological molecule should also be considered.
  • the solution is kept as low as possible in order to slow down the exchange between the bound xenon and free xenon, without broadening the NMR signals too much.
  • the solution is made viscous due to the use of one viscous solvent or the use of a suitable combination of solvents.
  • the viscosity of the solvent is preferably within the range of 500 mPs to 5000 mPs, more preferably within the range of 700 mPs to 1500 mPs.
  • the pressure of xenon is as high as possible, preferably higher than 5 xl0 5 N/m 2 (5 bar), more preferably higher than 5 x 10 6 N/m 2 (50 bar), even more preferably higher than 1 xl0 7 N/m 2 (100 bar) and particularly higher than 2xl0 7 N/m 2 (200 bar).
  • the pressure must never be so high that the biological molecule will be adversely effected.
  • Hype ⁇ olarised Xe is generated by optical pumping as described by B.D ⁇ ehuys et al., Appl.Phys.Lett. 69 (12), 1996.
  • the Isotopic composition of the gas is 80% 129 Xe and 0.25% Xe (the rest non-magnetic isotopes of Xe).
  • the degree of polarisation is estimated to be 10%.
  • Lyzozyme (28 mg) is dissolved in a mixture of D 2 O and methanol-cU (40:60) (3 ml) in a heavy- walled 10 mm NMR-tube. This mixture is subjected to four freeze-pump-thaw cycles of degassing.
  • the tube is then connected to the outlet of the polariser and frozen in liquid nitrogen.
  • the hype ⁇ olarized gas is generated and collected on a cold finger at liquid nitrogen temperature in a holding field of 200 mT over a period of 15 minutes which is estimated to give a volume of 50 ml of Xe at NTP.
  • a narrow Dewar vessel with liquid nitrogen is placed in a magnet with a field strength of 0.3 T.
  • the collected xenon is thawed and then refrozen in the NMR-tube in the 0.3 T magnet.
  • the sample tube is flame-sealed and the frozen sample is moved to the fringe field of the magnet of an NMR-spectrometer.
  • the NMR-spectrometer sample space is kept at a temperature of 293 K.
  • the sample is removed from the transport magnet and thawed by heating with the hand (protected from the cold) while standing as close to the NMR-magnet as possible.
  • the sample starts to thaw it is shaken vigorously and inserted into the spectrometer.
  • a 129 Xe spectrum is recorded and apart from the large peak due to the bulk xenon, a small peak, with a line width of 160 Hz, due to bound xenon can be observed at -158 ppm relative to bulk xenon.

Abstract

The invention relates to an in vitro method which comprises labelling a biological molecule with hyperpolarised xenon, and observing a magnetic resonance spectrum and/or image of the hyperpolarised xenon in the environment of the biological molecule. The spectrum/image provides information about the environment(s) at which atoms of xenon are bound to the biological molecule.

Description

NMR SPECTROSCOPY METHOD
This invention is concerned with nuclear magnetic resonance (NMR) spectroscopy and/or 5 NMR imaging. The technique involves observing the spectrum/image of a NMR active nuclear species in vitro in order to obtain information about the environment in which the species is present. The spectra of NMR active nuclei vary depending on their environment, and this is reported in the literature (PNAS, 93,12932-6, 1996).
10 Noble gases having non-zero nuclear spin can be hyperpolarised, i.e. have their polarisation enhanced over the equilibrium polarisation, e.g. by the use of circularly polarised light. Preferred techniques for hyperpolarisation include spin exchange with an optically pumped alkali metal vapour and metastability exchange. Noble gases to which this technique can be applied include 3He and 129Xe. As described by M S Albert et al in US Patent 5,545,396, the
15 technique can be used to prepare hyperpolarised noble gases that can be administered by inhalation for magnetic resonance imaging of the human body.
Xenon is chemically inert and has hydrophobic properties, and is capable of being weakly bound by hydrophobic regions of biological molecules (PNAS, 78, No 8, 4946-9, August 20 1981; Abstracts of the 11 Λ Annual Meeting of the Society for Magnetic Resonance in Medicine (1992) page 2104). Thus it is possible to "label" biological molecules with xenon.
This invention concerns the method of labelling biological molecules with hyperpolarised 129Xe. All macromolecules have a number of discrete hydrophobic and hydrophilic sites. Xenon 25 will bind by hydrophobic interactions to hydrophobic sites with different affinity. The xenon labels the biological compound by becoming weakly bound to it, e.g. at specific hydrophobic sites on a surface of or within a cavity of a protein or other macromolecule.
The NMR sensitivity of hyperpolarised xenon is highly increased compared to non- 30 hyperpolarised xenon. Another advantage of the present invention is the reversible and nondestructive nature of the bond between the xenon and the biological molecule. A further advantage is that the forming of the "bond" and subsequent measurement may be repeated if needed. In addition, since xenon is a gas (condensation temperature of -106°C), it and may easily and rapidly be separated from the biological molecule if necessary. Moreover, xenon is essential chemically inert and will not adversely effect the biological molecule.
One embodiment of the invention thus provides an in vitro method which comprises labelling a biological molecule with hyperpolarised xenon, and observing a magnetic resonance spectrum and/or image of the hyperpolarised xenon in the environment of the biological molecule. The spectrum/image provides information about the environment(s) at which atoms of xenon are bound to the biological molecule. Any conformational change of the biological molecule resulting e.g. from the binding (or the disappearance) of a ligand (e.g. a lipid, carbohydrate, peptide, polypeptide, nucleic acid or any sort of drug) or cleavage by an enzyme, will cause an alteration in the xenon NMR spectrum. Each hydrophobic site in the biological molecule may give rise to a specific and characteristic NMR shift.
A further embodiment of the present invention is to take NMR "fmgerprint(s)" of a known biological molecule. These fingerprints can subsequently be used to identify unknowns by direct comparison in a manner similar to infra-red spectroscopy.
A biological molecule as defined by the present invention is a monomeric or polymeric molecule that is present in biological systems or that is artificially introduced and is biologically active in such systems. Biological molecules include lipids, sugars and polysaccharides, nucleic acids (DNA, RNA), nucleosides, oligonucleosides, polynucleosides, nucleotides, oligonucleotides, polynucleotides, enzymes, vitamins and particularly peptides, polypeptides and proteins.
In one preferred embodiment of the invention, the labelled biological molecule is an assay reagent taking part in an assay method and wherein the assay reagent is labelled with hyperpolarised xenon. The labelling of the biological molecule with hyperpolarised xenon may be performed before, during or after performance of the assay. ___.
An assay method according to the present invention is a test involving a reaction of one or more biological molecules. The assays include for example competition assays (e.g. receptor- ligand antagonism, enzyme-substrate inhibitors, protein-protein interaction inhibitors), binding assays (e.g. receptor-ligand agonism, enzyme-substrate reactions, protein-protein interactions), immunoassays (e.g. for specific analytes), hybridisation assays (e.g. nuclease assays, mutation analysis, mRNA and DNA detection), test involving cells, organs and/or whole organisms. These tests may involve e.g. one or more lipids, saccharides, polynucleotides, oligonucleotides, nucleotides, peptides or proteins. Assays include binding studies performed on eukaryotic and prokaryotic microorganisms; binding studies performed on tissue in vitro; and binding studies in which an assay reagent is administered in vivo and an excretion product (e.g. urine, faeces, or breath) analysed by NMR in vitro.
By observing a change with time using NMR, the progress of a reaction can be followed during the course of an assay. Assays performed in vitro may conveniently be in multiwell plates, with either an assay reagent in the wells of the plate being labelled with hyperpolarised xenon, or a reagent being so labelled in bulk prior to being dispensed into individual wells of the plate.
Generally the biological molecule is present in a liquid medium into which the xenon is introduced as a gas. This may be achieved e.g. by bubbling it through the fluid or by contact with the biological molecule as a solid. Alternatively the xenon is introduced as a solution in a solvent, which is compatible with the biological molecule (e.g. in a lipophilic solvent such as a lipid or a fluorocarbon solvent).
The liquid medium used according to one embodiment of the present invention may be deuterated water, deuterated buffers or solvents, e.g. lipophilic solvents which may contain lipid bicelles, lipid vesicles, liposomes, cryptophanes and/or cyclodextrins.
129Xe has a natural abundance of 26.4%. The xenon used for this invention may be either the naturally occurring material or one artificially enriched in 12 Xe. A preferred degree of enrichment 129Xe is 40 % or more. A more preferred degree is 50 % or more and an even more preferred degree is 75 % or more. A particularly preferred degree of enrichment is 90 % or more. Bulk supplies of xenon enriched in 129Xe and hyperpolarised to a high degree are now available commercially and have a half life long enough to permit transport over substantial distances. While the half life of hyperpolarised I29Xe in the biological environments contemplated in this invention will be lower, it is expected to be amply sufficient to permit the desired spectra to be obtained. A preferred degree of hypeφolarisation is 8 % or more. A more preferred hypeφolarisation degree is 20 % or more and an even more preferred degree is 30 % or more. Ideally, the degree will approach 100 %.
In one embodiment of the invention, the temperature at the time xenon is added is above the temperature at which the biological molecule is frozen, but below the temperature at which the biological molecule may be denatured. Alternatively, xenon may be added to the frozen biological molecule, followed by thawing. However, the right temperature to achieve the optimal function of the biological molecule should also be considered.
In one embodiment of the invention, the solution is kept as low as possible in order to slow down the exchange between the bound xenon and free xenon, without broadening the NMR signals too much.
In a further embodiment of the invention, the solution is made viscous due to the use of one viscous solvent or the use of a suitable combination of solvents. The viscosity of the solvent is preferably within the range of 500 mPs to 5000 mPs, more preferably within the range of 700 mPs to 1500 mPs.
In one embodiment of the invention, the pressure of xenon is as high as possible, preferably higher than 5 xl05N/m2 (5 bar), more preferably higher than 5 x 106N/m2 (50 bar), even more preferably higher than 1 xl07N/m2 (100 bar) and particularly higher than 2xl07N/m2 (200 bar). However, the pressure must never be so high that the biological molecule will be adversely effected.
The invention is illustrated with reference to the following non-limiting Example.
Hypeφolarised Xe is generated by optical pumping as described by B.Dπehuys et al., Appl.Phys.Lett. 69 (12), 1996. The Isotopic composition of the gas is 80% 129Xe and 0.25% Xe (the rest non-magnetic isotopes of Xe). The degree of polarisation is estimated to be 10%. Lyzozyme (28 mg) is dissolved in a mixture of D2O and methanol-cU (40:60) (3 ml) in a heavy- walled 10 mm NMR-tube. This mixture is subjected to four freeze-pump-thaw cycles of degassing. The tube is then connected to the outlet of the polariser and frozen in liquid nitrogen. The hypeφolarized gas is generated and collected on a cold finger at liquid nitrogen temperature in a holding field of 200 mT over a period of 15 minutes which is estimated to give a volume of 50 ml of Xe at NTP. A narrow Dewar vessel with liquid nitrogen is placed in a magnet with a field strength of 0.3 T. The collected xenon is thawed and then refrozen in the NMR-tube in the 0.3 T magnet. The sample tube is flame-sealed and the frozen sample is moved to the fringe field of the magnet of an NMR-spectrometer. The NMR-spectrometer sample space is kept at a temperature of 293 K. The sample is removed from the transport magnet and thawed by heating with the hand (protected from the cold) while standing as close to the NMR-magnet as possible. When the sample starts to thaw it is shaken vigorously and inserted into the spectrometer. A 129Xe spectrum is recorded and apart from the large peak due to the bulk xenon, a small peak, with a line width of 160 Hz, due to bound xenon can be observed at -158 ppm relative to bulk xenon.

Claims

C L A I M S
1. An in vitro method which comprises labelling a biological molecule with hypeφolarised
190
Xe, and observing a magnetic resonance (NMR) spectrum and/or NMR image of the
190 hypeφolarised Xe in the environment of the biological molecule.
2. The method of claim 1 wherein the biological molecule is an assay reagent taking part in an assay method.
3. The method of claim 2 wherein the assay is a competition assay or an immunoassay .
4. The method of claim 2 wherein the assay is a hybridisation assay or a binding assay.
5. The method of any of claims 1 to 4 wherein the biological molecule is a peptide or a protein.
6. The method of any of claims 1 to 5 wherein the hypeφolarised 129Xe is enriched at a level of 40 % or more.
7. The method of any of claims 1 to 6 wherein the degree of hypeφolarisation is 8 % or more.
8. The method of any of claims 1 to 7 which is performed in a solution wherein the solvent has a viscosity in the range of 700 to 1500 mPs.
9. The method of any of claims 1 to 8 wherein the pressure of the xenon gas is at least 5 bar.
EP99963638A 1998-12-30 1999-12-23 Nmr spectroscopy method Withdrawn EP1141724A1 (en)

Applications Claiming Priority (3)

Application Number Priority Date Filing Date Title
GB9828853 1998-12-30
GBGB9828853.3A GB9828853D0 (en) 1998-12-30 1998-12-30 Nmr spectroscopy method
PCT/GB1999/004395 WO2000040972A1 (en) 1998-12-30 1999-12-23 Nmr spectroscopy method

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EP1141724A1 true EP1141724A1 (en) 2001-10-10

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EP (1) EP1141724A1 (en)
JP (1) JP2002534683A (en)
AU (1) AU1988000A (en)
GB (1) GB9828853D0 (en)
WO (1) WO2000040972A1 (en)

Families Citing this family (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6648130B1 (en) 1999-08-11 2003-11-18 Medi-Physics, Inc. Hyperpolarized gas transport and storage devices and associated transport and storage methods using permanent magnets
US6630126B2 (en) 2000-03-13 2003-10-07 Medi-Physics, Inc. Diagnostic procedures using direct injection of gaseous hyperpolarized 129Xe and associated systems and products
EP1324690A1 (en) 2000-07-13 2003-07-09 Medi-Physics, Inc. DIAGNOSTIC PROCEDURES USING ?129 XE SPECTROSCOPY CHARACTERISTIC CHEMICAL SHIFT TO DETECT PATHOLOGY i IN VIVO /i
US7179450B2 (en) 2001-09-20 2007-02-20 Medi-Physics, Inc. Methods for in vivo evaluation of pulmonary physiology and/or function using NMR signals of polarized Xe
NO20025738D0 (en) * 2002-11-29 2002-11-29 Amersham Health As Method

Family Cites Families (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5357959A (en) * 1993-04-16 1994-10-25 Praxair Technology, Inc. Altered dipole moment magnetic resonance imaging method
US5545396A (en) * 1994-04-08 1996-08-13 The Research Foundation Of State University Of New York Magnetic resonance imaging using hyperpolarized noble gases
RU2186405C2 (en) * 1996-03-29 2002-07-27 Лоренс Беркли Нэшнл Лэборэтори Amplification of nuclear magnetic resonance and magnetic resonance visualization in presence of hyperpolarized noble gases
ATE256293T1 (en) * 1997-01-08 2003-12-15 Amersham Health As METHOD FOR GENERATING IMAGE WITH MAGNETIC RESONANCE

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Title
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AU1988000A (en) 2000-07-24
WO2000040972A1 (en) 2000-07-13
JP2002534683A (en) 2002-10-15

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