Identification of pathogens in body fluids
Identification of infectious pathogens, particularly viruses, bacteria and other microorganisms is effected with a method whereby pathogens of acute infections can be identified, without first culturing them in external nutrient media, by mass spectrometric measurement of their protein profiles obtained from pathogens directly precipitated from body fluid into pellets by centrifuging. With this method, pathogens which cause acute infections can be identified in less than one hour.
1. A method for the mass spectrometric identification of pathogens in a blood culture, the method comprising the steps of:
a) destroying blood cells in the blood culture;
b) precipitating the pathogens into a pellet;
c) subjecting the pathogens to a mass spectrometric analysis of their proteins; and
d) identifying the pathogens by comparison of protein mass spectra with reference protein mass spectra.
2. The method according to claim 1 , wherein, in step b), the pathogens in the pellet are smeared directly onto a mass spectrometric sample support plate and sprinkled with matrix solution for ionization of proteins thereof by matrix-assisted laser desorption.
3. The method according to claim 1 , wherein step b) comprises precipitating by centrifuging.
4. The method according to claim 3 , wherein, in step b), proteins are extracted from the pathogens while the pellet with the pathogens still rests in a centrifuge tube and an extraction fluid is subjected to mass spectrometric measurement of the proteins.
5. The method according to claim 4 , wherein, for extraction of the proteins, the pathogens of the pellet are decomposed by chemical, physico-chemical or physical means.
6. The method according to claim 4 , wherein the extraction fluid is subjected to mass spectrometric analysis using electrospray ionization.
7. The method according to claim 5 , wherein the extraction fluid is subjected to mass spectrometric analysis using electrospray ionization.
8. The method according to claim 4 , wherein the extraction fluid is subjected to mass spectrometric analysis using ionization by matrix-assisted laser desorption.
9. The method according to claim 5 , wherein the extraction fluid is subjected to mass spectrometric analysis using ionization by matrix-assisted laser desorption.
10. The method according to claim 8 , wherein the extraction fluid is applied to a mass spectrometric sample support plate onto which a layer of matrix crystals has already been deposited.
11. The method according to claim 8 , wherein a matrix substance solution is added to the extraction fluid before this is applied to the sample support plate.
12. A method for the mass spectrometric identification of pathogens in a blood culture, the method comprising the steps of:
a) precipitating the pathogens together with blood cells into a pellet;
b) destroying the blood cells in the pellet;
c) subjecting the pathogens to a mass spectrometric analysis of their proteins; and
d) identifying the pathogens by comparison of protein mass spectra with reference protein mass spectra.
13. The method according to claim 12 , wherein the blood cells which are present in the pellet are destroyed by osmosis.
14. The method according to claim 13 , wherein distilled water is used for destroying the blood cells in the pellet by osmosis.
15. The method according to claim 12 , wherein step a) comprises precipitating by centrifuging.
16. The method according to claim 15 , wherein washing and a further centrifuging between steps b) and c) deposits a pellet containing enriched pathogens.