Method and device for analyzing the intracellular chemical state of living cells by nuclear magnetic resonance
AssigneeBruker BioSpin GmbH
InventorsBenjamin Gonzalez, Martial Piotto, J. Gaspard Huber
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Abstract
The invention concerns a method for analysing the intracellular chemical state of living cells by nuclear magnetic resonance (NMR). Said method comprises a step which consists in preparing the cell sample and a step which consists in an NMR analysis of the sample, both steps being carried out at very low temperature so as to freeze the biochemical state of the living cells to obtain specific and accurate reproducible measurements of the cellular functioning in given conditions. The invention also concerns the device coupled to the NMR spectrometer for implementing the method of analysis.
Claims15
A method for preparing a sample and for analyzing the chemical state of living cells by nuclear magnetic resonance (NMR) comprising:
- i) preparing a sample of living cells so as to set the chemical state of the cells by:
- a) subjecting said sample of living cells to a temperature at or lower than −80° C. so as to stop intracellular reactions, by directly immersing said sample in liquid nitrogen, followed by,
- b) lyophilizing said sample;
- ii) storing said sample at said temperature less than or equal to −80° C.
- iii) just before performing the NMR analysis, mixing and redissolving said lyophilized sample, at a temperature between 0° C. and −20° C., with a solvent having a very low melting point and being liquid at the temperature between 0° C. and −20° C.;
- iv) obtaining, at the temperature between 0° C. and −20° C., at least one NMR spectrum and/or at least one NMR measurement value through High Resolution Magic Angle Spinninig (HRMAS) NMR;
- and
- v) comparing said at least one HRMAS NMR spectrum and/or at least one HRMAS NMR measurement value with at least one reference NMR spectrum and/or at least one reference NMR measurement value obtained on at least one reference sample of living cells.
A method for preparing a sample and for analyzing the chemical state of living cells by nuclear magnetic resonance (NMR) comprising:
- i) preparing a sample of living cells so as to set the chemical state of the cells by:
- a) subjecting said sample of living cells to a temperature at or lower than −80° C. so as to stop intracellular reactions, by directly immersing said sample in liquid nitrogen, followed by,
- b) lyophilizing said sample;
- ii) storing said sample at said temperature less than or equal to −80° C.;
- iii) just before performing the NMR analysis, mixing and redissolving said lyophilized sample, at a temperature between 0° C. and −20° C., with a solvent having a very low melting point and being liquid at the temperature between 0° C. and −20° C.;
- iv) obtaining, at the temperature between 0° C. and −20° C., and without further extraction of metabolites, at least one NMR spectrum and/or at least one NMR measurement value through High Resolution Magic Angle Spinninig (HRMAS) NMR;
- and
- v) comparing said at least one HRMAS NMR spectrum and/or at least one HRMAS NMR measurement value with at least one reference NMR spectrum and/or at least one reference NMR measurement value obtained on at least one reference sample of living cells.
Dependencies are inferred from the claim text.