Method for detecting and identifying microorganism causative of infection
AssigneesFuso Pharmaceutical Industries, Ltd., Tsuneya Ohno
InventorsTsuneya Ohno, Akio Matsuhisa
Life of this patent
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Abstract
Causative microorganisms of infectious diseases are detected and/or identified rapidly and high-sensitively by taking phagocytes from the clinical specimens containing active phagocytes, immobilizing the phagocytes so taken, treating the phagocytes to improve cell membrane permeabilities thereof, further treating the phagocytes to bare DNA in the causative microorganisms which might be existed in the phagocytes, and detecting the causative microorganisms with DNA probes which can hybridize with such DNA under stringent conditions.
Claims20
1-18. (canceled)
A method for detecting and/or identifying microorganisms causative of infectious diseases, comprising:
- immobilizing phagocytes from a clinical specimen containing active phagocytes that may contain microorganisms causative of infectious diseases;
- treating the phagocytes to increase phagocyte cell membrane permeability;
- further treating the phagocytes to bare DNA in microorganisms that may exist in the phagocytes;
- in situ hybridizing the bared DNA with at least one DNA probe;
- and
- detecting and/or identifying the causative microorganisms by detecting hybridization of at least one probe to the bared DNA,
- wherein the method comprises at least one condition selected from the group consisting of:
- (1) the phagocytes in the immobilizing step have a cell density between 5×106 cells/ml and 1×108 cells/ml;
- (2) applying from about 1 Unit/ml to about 1,000 Units/ml of Lysostaphin to the phagocytes in the further treating step, to bare the DNA;
- (3) applying from about 1,000 Units/ml to about 1,000,000 Units/ml of Lysozyme to the phagocytes in the further treating step to bare the DNA;
- (4) applying from about 10 Units/ml to about 10,000 Units/ml of N-acetylmuramidase to the phagocytes in the further treating step to bare the DNA;
- (5) applying from about 50 Units/ml to about 500 Units/ml of Zymolyase to the phagocytes in the further treating step to bare the DNA;
- (6) applying surfactant in the step of in situ hybridization;
- (7) the at least one DNA probe is from about 350 to about 600 bases in length;
- and
- (8) the concentration of the at least one DNA probe is from about 0.1 ng/μl to about 2.2 ng/μl.
A kit for detecting and/or identifying microorganisms causative of infectious diseases from phagocytes, comprising:
- (1) a composition comprising at least one enzyme selected from the group consisting of Lysostaphin, Lysozyme, N-acetylmuramidase and Zymolyase;
- and
- (2) at least one DNA probe for detecting DNA of a microorganism.
Dependencies are inferred from the claim text.