Humidity stabilization during the preparation of biological samples for spectrometry
The invention proposes preparing biological samples for spectrometry which contain cell structures and/or whole cells of human or animal origin (e.g. thin human and animal tissue sections) or prokaryotes (e.g. microorganisms), and which require constant relative humidity, in a temperature-controlled gas volume whose humidity is determined by a saturated substance solution, for example a suitable salt solution. The invention exploits a physico-chemical phenomenon called “deliquescence”, which manifests itself by keeping the relative humidity above the saturated substance solution constant with a high degree of precision when a specified temperature is maintained. Pure succinic acid exhibits deliquescence at approx. 99% relative humidity, for example. Since an enormous variety of deliquescent salts and other suitable substances are available, it is possible to find the suitable substance for almost any desired relative humidity, with adjustment of the temperature, where necessary.
1. A method for the preparation of a biological sample of human or animal origin for spectrometric analysis, comprising:
providing the biological sample of human or animal origin;
subjecting the biological sample to a preparation which renders it fit for a following spectrometric analysis, wherein the preparation takes place in a gas volume in which the relative humidity is kept constant by deliquescence; and
transmitting the prepared biological sample to spectrometric analysis.
2. The method according to claim 1 , wherein the biological sample is a thin tissue section and the spectrometric analysis is imaging mass spectrometry.
3. The method according to claim 2 , wherein the imaging mass spectrometry works with ionization by matrix-assisted laser desorption in a time-of-flight mass spectrometer.
4. The method according to claim 2 , wherein the preparation comprises chemical or enzymatic conversions of molecules in the thin tissue section, where a set level of humidity causes the tissue to swell to the desired extent, which is advantageous for the chemical or enzymatic reactions while largely maintaining the positional accuracy of the molecules.
5. The method according to claim 4 , wherein the swelling of the tissue is assisted by a hygroscopic substance which is applied in a measured quantity.
6. The method according to claim 2 , wherein the tissue is provided in chemically fixed form, for example fixed by formaldehyde.
7. The method according to claim 6 , wherein the tissue is provided embedded in an organic solid material, such as paraffin.
8. The method according to claim 2 , wherein the tissue is provided in frozen form.
9. The method according to claim 1 , wherein macromolecules of the biological sample are broken into fragments by enzymatic digestion.
10. The method according to claim 1 , wherein, during preparation, the temperature of the biological sample is briefly reduced to below the dew point to facilitate the controlled formation of a cohesive film of moisture on the surface of the sample.
11. The method according to claim 1 , further comprising setting a high level of humidity just below the dew point.
12. The method according to claim 11 , where the relative humidity is set to be equal to or higher than 95%.
13. The method according to claim 1 , wherein the preparation of the biological sample takes place directly on the same sample support which subsequently serves as substrate for a mass spectrometric or infrared-spectrometric measurement.
14. The method according to claim 13 , wherein the sample support takes the form of a flat steel plate, or flat ceramic plate/glass plate having a conductive coating.
15. The method according to claim 1 , wherein the gas volume is supplied with a gas flow which has been brought to a specified relative humidity by interaction with a saturated solution of a deliquescent substance.
16. A preparation chamber to carry out a method according to claim 1 , which comprises a gas-tight chamber, a holder for the sample support, a device for controlling the temperature of the chamber, and a dish for a saturated solution and a deliquescent substance to be dissolved.
17. The preparation chamber according to claim 16 , further comprising a lock for the introduction and removal of the sample support.
18. The preparation chamber according to claim 16 , further comprising a device for gently agitating the saturated solution and/or a device for gently agitating gas in the chamber.
19. A The method according to claim 1 , wherein the biological sample contains cell structures and/or whole cells of human or animal origin.
20. A method for the preparation of a prokaryotic or fungi biological sample for a spectrometric analysis, comprising:
providing the prokaryotic or fungi biological sample;
subjecting the prokaryotic of fungi biological sample to a preparation which renders it fit for a following spectrometric analysis, wherein the preparation takes place in a gas volume in which the relative humidity is kept constant by deliquescence; and
transmitting the prepared prokaryotic or fungi biological sample to spectrometric analysis.
21. The method according to claim 20 , wherein the preparation of the prokaryotic or fungi biological sample is carried out on living microbes, which are incubated on a sample support plate in droplets of a nutrient liquid.
22. The method according to claim 21 , wherein the droplets contain a measured quantity of an anti-microbial substance in order to allow for susceptibility testing.
23. The method according to claim 20 , wherein the preparation of the prokaryotic or fungi biological sample is followed by infrared spectrometric analysis, whose aim is to determine a subspecies or variety of the sample.
24. The method according to claim 20 , wherein the relative humidity is kept at about between 10% and 30%.