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1. WO2011063389 - NORMALISATION DE BIOMARQUEURS DE PLAQUETTES

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CLAIMS

1. A method for assessing a biomarker level for a platelet preparation, the method

comprising:

(a) determining the level of a surrogate marker in a platelet preparation sample, wherein the surrogate marker corresponds to platelet number, platelet concentration or platelet volume;

(b) determining the level of a biomarker in the sample,

(c) normalizing the level of the biomarker in the sample to the level of the surrogate marker, whereby a normalized biomarker for the sample is determined.

2. The method of claim 1, wherein the normalizing step comprises dividing the value obtained for the level of the biomarker in the sample by the value obtained for the level of the surrogate marker.

3. The method of claim 1, wherein the surrogate marker is polymerized or monomeric actin.

4. The method of claim 1, wherein step (a) comprises placing the sample obtained from an individual under conditions that induce actin polymerization, such that actin in the sample is substantially polymerized.

5. The method of claim 5, wherein a high concentration of salt promotes actin polymerization.

6. The method of claim 1, wherein the biomarker is an angiogenic regulator.

7. A method for identifying a surrogate marker for platelet number, platelet concentration, or platelet volume, the method comprising:

(a) assaying the amount of a plurality of candidate markers in each sample of a series of samples prepared from a single platelet preparation according to a sampling factor;

(b) comparing the amount of each candidate marker in each sample to the amount of candidate marker predicted according to the sampling factor, wherein the comparing step identifies the candidate marker of the plurality assayed in step (a) that has the closest correlation between the amount of candidate marker predicted and the amount of candidate

marker measured, whereby the candidate marker is identified as a surrogate marker for platelet number, platelet concentration, or platelet volume.

8. The method of claim 6, wherein said platelet preparation comprises lysed platelets.

9. The method of claim 6, further comprising testing the identified surrogate marker for variation under different physiological conditions.

10. A method for normalizing the amount of a biomarker in a sample, the method comprising normalizing the amount of a biomarker measured in a platelet preparation relative to a surrogate marker identified using the method of claim 6.

11. The method of claim 10, wherein the biomarker is an angiogenic protein.

12. The method of claim 8, wherein the platelet preparation is obtained from a patient sample.

13. The method of claim 10, further comprising comparing a normalized level of the biomarker to a reference level to detect a change in the level of the biomarker in the patient sample.

14. A method for assessing a biomarker level for a platelet preparation, the method comprising:

(a) placing a sample of isolated platelets obtained from said individual under conditions that induce actin polymerization, such that actin in said sample is substantially polymerized;

(b) contacting said sample with an agent that selectively binds polymerized actin and detecting formation of a complex between said agent and polymerized actin, whereby the level of actin in said sample is measured;

(c) measuring the level of a biomarker in said sample;

(d) normalizing the level of said biomarker in said sample to the measured level of polymerized actin in said sample, whereby a normalized biomarker for the sample is determined.

15. A method for assessing a change in biomarker level of an individual, the method comprising:

(a) placing a sample of isolated platelets obtained from said individual under conditions that induce actin polymerization, such that actin in said sample is substantially polymerized;

(b) contacting said sample with an agent that selectively binds polymerized actin and detecting formation of a complex between said agent and polymerized actin, whereby the level of actin in said sample is measured;

(c) measuring the level of a biomarker in said sample;

(d) normalizing the level of said biomarker in said sample to the measured level of polymerized actin in said sample,

(e) comparing a normalized level of said biomarker in said sample to a reference, wherein a difference in the normalized level of said biomarker compared to said reference indicates a change in the level of said biomarker of said individual.

16. The method of claim 15, wherein said conditions that induce actin polymerization comprise a high concentration of salt.

17. The method of claim 15, wherein the biomarker is an angiogenic regulator.

18. The method of claim 17, wherein a change in the level of the angiogenic regulator is indicative of a change in angiogenic state and/or the presence of an angiogenic disorder.

19. The method of claim 18, wherein said angiogenic disorder is the presence of a tumor-associated disease.

20. The method of claim 15, wherein said reference is obtained from biological samples obtained from a population of individuals.

21. The method of claim 20, wherein each individual of said population is free from an angiogenic disorder.

22. The method of claim 12, wherein said reference is obtained from isolated platelets obtained from said individual at an earlier time point.

23. A kit for detecting a normalized level of at least one biomarker in platelets, the kit comprising:

(a) at least one reagent which, when contacted with an isolated platelet sample induces actin polymerization or depolymerization; and

(b) an agent that selectively binds either polymerized actin where the reagent of step (a) induces actin polymerization, or monomeric actin where the reagent of step (a) induces actin depolymerization;

(c) an agent that binds a biomarker; and

(d) packing materials and instructions for normalizing the level of the at least one biomarker to the level of polymerized or monomeric actin.

24. The kit of claim 23, further comprising a solid support.

25. The kit of claim 23, further comprising a reagent that generates a detectable signal.

26. The kit of claim 23, further comprising a polymerized actin positive control.

27. The kit of claim 23, further comprising an agent that binds at least one other biomarker.

28. A computer readable storage medium having computer readable instructions recorded thereon to define software modules for implementing on a computer a method for assessing a biomarker level in a platelet sample, said computer readable storage medium comprising:

(a) instructions for storing and accessing data representing a level of a biomarker and a level of a surrogate marker determined for a sample of isolated platelets obtained from at least one individual;

(b) instructions for normalizing said level of said biomarker to said level of said surrogate marker via a normalization module, thereby producing a normalized level of said biomarker,

(c) instructions for displaying retrieved content to a user, wherein the retrieved content comprises a normalized biomarker level.

29. The computer readable storage medium of claim 27, further comprising instructions for comparing said normalized level of said biomarker to reference data stored on said storage device using a comparison module, whereby a change in the biomarker level is determined.

30. The method of claim 28, wherein the surrogate marker is polymerized or monomeric actin.

31. A computer system for obtaining data from a sample of isolated platelets obtained from at least one individual, the system comprising:

(a) a specimen container to hold said sample;

(b) a determination module configured to determine read-out information, wherein said read-out information comprises

1) information representing an amount of a surrogate marker of platelet number, platelet concentration or platelet volume, and

2) information representing an amount of a biomarker measured in the sample,

(c) a storage device configured to store data output from said determination module,

(d) a normalization module configured to normalize information representing a level of said biomarker to information representing an amount of said surrogate marker;

(e) a display module for displaying retrieved content to the user, wherein the retrieved content comprises a normalized biomarker level.

32. The computer system of claim 31, further comprising a comparison module adapted to compare the data obtained from said normalization module with reference data on said storage device, whereby a change in the level of said biomarker is determined.

33. The method of claim 32, wherein the surrogate marker is polymerized or monomeric actin.