MASS SPECTROMETRIC DIAGNOSIS OF SEPTICEMIA
The invention mainly relates to the mass spectrometric identification of pathogens in blood cultures from bloodstream infections (septicemia). The invention provides a method with which microbial pathogens can be separated in purified form from blood after a relatively brief cultivation in a blood culture flask, without any interfering human proteins or any residual fractions of blood particles such as erythrocytes and leukocytes, and can be directly identified by mass spectrometric measurement of their protein profiles. The method is based on the use of relatively strong tensides to destroy the blood particles by dissolving the weak cell membranes and most of the internal structures of the blood particles; in spite of the fact that tensides are regarded as strong ionization inhibitors in MALDI and other ionization processes required for mass spectrometric measurements. This method allows unknown pathogens to be obtained in their pure form by centrifuging or filtration and to be identified on the taxonomic level of species or subspecies. Problems with DNA from high levels of leukocytes can be resolved by special measures. After sufficient cultivation, the identification in a mass spectrometric laboratory takes only half an hour.
1 . An analysis kit for mass spectrometric identification of microbes of a blood sample of a blood culture, the analysis kit comprising:
a tenside solution comprising a defoamer;
the tenside solution prepared for dissolving blood particles of the blood sample of the blood culture prior to deposition of the microbes by centrifuging or filtration, wherein the microbes comprise bacteria and yeast; and
wherein the tenside solution enables the microbes to be identified by acquiring a mass spectrum of the deposited microbes.
2 . The analysis kit of claim 1 , further comprising instructions describing a mass spectrometric method for using the tenside solution to identify microbes in the blood sample.
3 . The analysis kit of claim 1 , further comprising a centrifuge tube in which the dissolved blood particles are to be centrifuged.
4 . The analysis kit of claim 1 , wherein the tenside comprises sodium dodecyl sulfate (SDS).
5 . The analysis kit of claim 1 , wherein the tenside solution comprises between about five percent and about twenty percent of the sodium dodecyl sulfate (SDS).
6 . The analysis kit of claim 1 , further comprising an anticoagulant for adding to a blood sample when preparing the blood culture.
7 . The analysis kit of claim 1 , further comprising one or more nucleases for added to the blood sample.
8 . The analysis kit of claim 1 , further comprising distilled water for diluting the blood sample by a dilution factor between about 1:2 and about 1:10.
9 . The analysis kit of claim 1 , further comprising distilled water for diluting the blood sample by a dilution factor of about 1:5.
10 . The analysis kit of claim 1 , wherein the deposited microbes are to be disintegrated by sonication or mechanical treatment.
11 . The analysis kit of claim 1 , further comprising a solution of formic acid or trifluoroethanoic acid and acetonitrile for disintegrating the deposited microbes.
12 . The analysis kit of claim 11 , further comprising:
a matrix substance for matrix assisted laser desorption/ionization; and
a mass spectrometric sample support on which a measurement sample is to be prepared using the matrix substance and the disintegrated microbes, wherein soluble proteins of the disintegrated microbes are embedded in crystals of the measurement sample.
13 . The analysis kit of claim 1 , further comprising:
a matrix for matrix assisted laser desorption/ionization.
14 . The analysis kit of claim 1 , further comprising:
a mass spectrometric sample support on which a measurement sample is to be prepared using the matrix substance and the deposited microbes.
15 . The analysis kit of claim 1 , wherein the deposited microbes are to be washed and re-deposited by centrifuging before acquiring the mass spectrum for identification.
16 . The analysis kit of claim 1 , wherein the microbes are to be identified by acquiring the mass spectrum of the deposited microbes using a time-of-flight mass spectrometer with ionization by matrix assisted laser desorption (MALDI).
17 . An analysis kit for mass spectrometric identification of microbes of a blood sample of a blood culture, the analysis kit comprising:
a tenside solution comprising a defoamer;
the tenside solution prepared for dissolving the blood particles of the blood sample of the blood culture prior to deposition of the microbes by sedimentation or micro-filtration, wherein the microbes comprise bacteria and yeast; and
wherein the tenside solution enables the microbes to be identified by acquiring a mass spectrum of the deposited microbes.