Method of identifying and assessing DNA euchromatin in biological cells for detecting disease, monitoring wellness, assessing bio-activity, and screening pharmacological agents

• US 7,153,691 B2
• Filed: 11/13/2002
• Issued: 12/26/2006
• Est. Priority Date: 11/13/2002
• Status: Expired due to Fees

• Current Assignee: G6 Science Corp.
• Original Assignee: G6 Science Corp.
• Inventors: Ferguson, Gary William
• Primary Examiner(s): WALLENHORST, MAUREEN
• Application Number:
  US10/293,609
• Publication Number:
  US 20040092026A1
• Time in Patent Office: 1,504 Days
• Field of Search: 436/63, 436/117, 436/164, 436/166, 436/17, 435/6
• US Class Current: 436/63
• CPC Class Codes: C12Q 1/68 involving nucleic acids
• CPC Class Codes: C12Q 1/6841 In situ hybridisation

Abstract

The present invention is a method of identifying and assessing DNA euchromatin in biological cells. The amount and/or distribution of DNA euchromatin generally relates to RNA/protein synthesis, which may change in certain conditions, such as disease, and/or cellular response to environmental and/or chemical agents. The present invention detects the potential presence of disease by assessing DNA euchromatin in ostensibly normal cells and similarly provides means to monitor and assess treatment response. The present invention may also be used to assess how application (or removal) of influences, such as environment, radiation, chemical agents, medications, herbs, vitamins, etc., interact to effect cells, establishing uses in monitoring wellness and pharmacological screening. Additional uses include assessing age related disorders, allergies, infections, Alzheimer'"'"'s disease, ‘Time of Death’, chronic fatigue syndrome, etc. The method may be advantageously applied to biological cells including micro-organisms, plants, animals, or cultured cells, alone or in conjunction with other markers.

Claims

1. A method of assessing euchromatin, comprising:

preferentially staining euchromatin in biological cells;

measuring at least one variable, said at least one variable comprising at least one of an amount of said euchromatin and a distribution of said euchromatin; and

comparing said at least one variable to a reference value, wherein the reference value represents at least one of a known amount of euchromatin and a known distribution of euchromatin in reference cells.

2. The method of claim 1, further comprising using an absorbance stain.

3. The method of claim 2, wherein said absorbance stain comprises at least one of pararosaniline, azure A, a thiazine derivative, and acriflavine.

4. The method of claim 1, further comprising using a fluorescence stain.

5. The method of claim 4, wherein said fluorescence stain comprises at least one of a propidium iodide, an adenine-thiamine selective stain, a green nucleic acid stain, and a cytosine-guanine stain.

6. The method of claim 1, further comprising using at least one of monoclonal antibody or a gene marker.

7. The method of claim 1, wherein said staining step comprises Feulgen staining.

8. The method of claim 7, wherein said Feulgen staining step ccmprises preferentially hydrolyzing said euchromatin with an acid having a concentration in the range of 0.1 to 5.0 N and performing said Feulgen staining step at less than 25 degrees Celsius.

9. The method of claim 8, wherein said hydrolyzing step is performed at approximately 15 degrees Celsius.

10. The method of claim 8, wherein said acid is hydrochloric acid.

11. The method of claim 8, wherein said hydrolyzing step is performed for one hour.

12. The method of claim 8, wherein said hydrolyzing step is performed for a time period between one minute and two hours.

13. The method of claim 8, wherein said hydrolyzing step is performed for a time period between 15 minutes and two hours.

14. The method of claim 1, further comprising assessing at least one of an amount of total DNA and a distribution of total DNA in the biological cells.

15. The method of claim 14, futher comprising computing a parameter comprising at least one of a ratio of said amount of euchromatin to said amount of total DNA and the ratio of said distribution of euchromatin to said distribution of total DNA.

16. The method of claim 15, wherein said parameter is computed for a subpopulation of the biological cells.

17. The method of claim 1, wherein said reference value is stored in a database.

18. The method of claim 1, wherein said reference value is determined from substantially healthy biological cells.

19. The method of claim 1, wherein said reference value is determined from diseased cells.

20. The method of claim 1, wherein said method is used for at least one of detecting disease, monitoring weilness, assessing bio-activity, and pharmacological screening.

21. A method of assessing euchromatin in biological cells, comprising:

preferentially staining euchromatin in a first set of biological cells taken from an organism;

measuring at least one variable, said at least one variable comprising at least one of an amount of said euchromatin and a distribution of said euchromatin in said first set of biological cells;

expressing at least one of said measured amount and said measured distributior in said first set of biological cells as a basal cell response factor relating to said euchromatin;

altering an influence on the organism;

preferentially staining euchromatin in a second set of biological cells taken from the organism after said altering step;

measuring at least one variable, said at least one variable comprising at least one of an amount of said euchromatin and a distribution of said euchromatin in said second set of biological cells;

expressing at least one of said measured amount and said measured distribution in said second set of biological cells as an exposure cell response factor relating to said euchromatin; and

comparing said basal cell response factor to said exposure cell response factor.

22. The method of claim 21, wherein said influence comprises at least one of an environment, radiation, a chemical agent a medication, an herb, and a vitamin.

23. The method of claim 21, wherein said altering step comprises reducing the organisms'"'"'s exposure to said influence.

24. The method of claim 21, wherein said altering step comprises increasing the organism'"'"'s exposure to said influence.

25. The method of claim 21, further comprising using an absorbance stain.

26. The method of claim 25, wherein said absorbance stain comprises at least one of pararosaniline, azure A, a thiazine derivative, and acriflavine.

27. The method of claim 21, further comprising using a fluorescence stain.

28. The method of claim 27, wherein said fluorescence stain comprises at least one of a propidium iodide, an adenine-thiamine selective stain, a green nucleic acid stain, and a cytosine-guanine stain.

29. The method of claim 21, further comprising using at least one of a monoclonal antibody or a gene marker.

30. The method of claim 21, wherein said staining step comprises Feulgen staining.

31. The method of claim 30, wherein said Feulgen staining step comprises preferentially hydrolyzing said euchromatin with an acid having a concentration in the range of 0.1 to 5.0 N and performing said Feulgen staining step at less than 25 degrees Celsius.

32. The method of claim 31, wherein said hydrolyzing step is performed at approximately 15 degrees Celsius.

33. The method of claim 31, wherein said acid is hydrochloric acid.

34. The method of claim 31, wherein said hydrolyzing step is performed for one hour.

35. The method of claim 31, wherein said hydrolyzing step is performed for a time period between one minute and two hours.

36. The method of claim 31, wherein said hydrolyzing step is performed for a time period between 15 minutes and two hours.

37. The method of claim 21, further comprising assessing at least one of an amount of total DNA and a distribution of total DNA in the biological cells.

38. The method of claim 37, further comprising computing a parameter comprising at least one of the ratio of said amount of euchromatin to said amount of total DNA and the ratio of said distribution of euchromadn to said distribution of total DNA.

39. The method of claim 38, wherein said parameter is computed for a subpopulation of the biological cells.

40. The method of claim 21, further comprising comparing at least one of said basal cell response factor and said exposure cell response factor to a reference value.

41. The method of claim 40, wherein said reference value is stored in a database.

42. The method of claim 40, wherein said reference value is determined from substantially healthy biological cells.

43. The method of claim 40, wherein said reference value is determined from diseased cells.

44. The method of claim 21, wherein said method is used for at least one of detecting disease, monitoring wellness, assessing bin-activity, and pharmacological screening.