Cytological stain composition

• US 6,593,102 B2
• Filed: 11/12/2001
• Issued: 07/15/2003
• Est. Priority Date: 10/29/1999
• Status: Expired due to Term

• Current Assignee: Cytyc Corporation (Hologic Incorporated)
• Original Assignee: Cytyc Corporation (Hologic Incorporated)
• Inventors: Lapen, Daniel C., Zahniser, David J., Isenstein, Louise M., Mui, Katherine K., Soule, Norman W.
• Primary Examiner(s): Leary, Louise N.
• Application Number:
  US10/054,578
• Publication Number:
  US 20020150967A1
• Time in Patent Office: 610 Days
• Field of Search: 435/40.5, 435/4, 356/399
• US Class Current: 435/40.5
• CPC Class Codes: G01N 1/30 Staining; Impregnating ; Fixation; Dehydration; Multistep processes for preparing samples of tissue, cell or nucleic acid material and the like for analysis
• CPC Class Codes: G01N 15/1433 using image recognition
• CPC Class Codes: G01N 15/1468 with spatial resolution of the texture or inner structure of the particle
• CPC Class Codes: G01N 15/147 the analysis being performed on a sample stream
• CPC Class Codes: G01N 2015/1006 for cytology
• CPC Class Codes: G01N 2015/1472 with colour
• CPC Class Codes: G01N 2015/1488 Methods for deciding
• CPC Class Codes: G01N 21/3151 using two sources of radiation of different wavelengths G01N21/33 - G01N21/39 take precedence
• CPC Class Codes: G06T 2207/30024 Cell structures in vitro; Tissue sections in vitro
• CPC Class Codes: G06T 7/0012 Biomedical image inspection

Abstract

The present invention relates to the analysis of cytological material. Specifically, the invention relates to stains and methods of producing the stains, methods of staining cells for cytological or histological analysis to contrast the nuclear portion of the cell from the cytoplasmic portion, and systems and methods for illuminating and imaging a cytological sample. The analysis can be automated or manual.

Claims

1. A cytological staining solution, comprising:

methanol;

a phenol derivative; and

thionin.

2. The staining solution of claim 1, wherein the methanol is various grades.

3. The staining solution of claim 1, wherein the phenol derivative is selected from the group consisting of phenol ethers, phenol amines, akylphenols, hydroxy benzene compounds, bisphenols, and halogenated phenolic compounds.

4. The staining solution of claim 1, wherein the phenol derivative is phenoxyethanol.

5. The staining solution of claim 1, wherein the thionin comprises a certified dye powder.

6. The staining solution of claim 1 further comprising an acid, wherein the acid is selected from the group consisting of acetic acid, phosphoric acid, nitric acid, formic acid, citric acid, sulfuric acid, and hydrochloric acid.

7. The staining solution of claim 1, wherein the acid is glacial acetic acid.

8. The staining solution of claim 1, wherein pH of the staining solution is acidic.

9. The staining solution of claim 1, wherein pH of the staining solution is between about 5 to about 7.

10. The staining solution of claim 1, wherein pH of the staining solution is about 6.7+/−

0.05.

11. The staining solution of claim 1, wherein the phenol derivative has a weight to volume ratio of about 0.8% to about 1.2%.

12. The staining solution of claim 1, wherein the phenol derivative has a weight to volume ratio of about 1%.

13. The staining solution of claim 1, wherein the thionin is synthesized.

14. The staining solution of claim 1, wherein the thionin has a weight to volume ratio of about 0.2% to about 0.4%.

15. The staining solution of claim 1, wherein the thionin has a weight to volume ratio of about 0.3%.

16. The staining solution of claim 1, wherein the thionin has a weight to volume ratio of about 0.345%.

17. A method of producing a cytological staining solution, the method comprising the steps of:

mixing methanol, a phenol derivative, and thionin;

stirring the mixture;

filtering the mixture; and

adjusting pH of the mixture to a value of about 6.7 by adding an acid thereto to produce the staining solution.

18. The staining solution of claim 17, wherein the phenol derivative is selected from the group consisting of phenol ethers, phenol amines, akylphenols, hydroxy benzene compounds, bisphenols, and halogenated phenolic compounds.

19. The staining solution of claim 17, wherein the phenol derivative is phenoxyethanol.

20. The method of claim 17, wherein the acid is chosen from the group consisting of acetic acid, phosphoric acid, nitric acid, formic acid, citric acid, sulfuric acid, and hydrochloric acid.

21. The method of claim 17, wherein the acid is glacial acetic acid.

22. The method of claim 17, wherein the mixture is mixed for at least about 1 hour.

23. The method of claim 17, further comprising the step of mixing the mixture while adding the acid.

24. The method of claim 17, wherein the mixing step comprises mixing an equivalent ratio of about one liter of methanol, about 10 grams of phenol derivative, and about 3.45 grams of thionin.

25. The method of claim 17, wherein the filtering step utilizes a filter with about 1-20 micron particle retention.

26. A method of staining cytological material, the method comprising staining the cytological material with a thionin-phenol derivative solution.

27. The method of claim 26, further comprising the step of rinsing the cytological material in a salt bath prior to staining.

28. The method of claim 26, further comprising the step of counterstaining the cytological material with a second stain.

29. The method of claim 27, wherein the salt bath is about a 10% salt solution.

30. The method of claim 26, wherein the thionin-phenol derivative solution is about a 0.3% thionin solution.

31. The method of claim 26, wherein the thionin-phenol derivative solution comprises methanol, a phenol derivative, and thionin.

32. The method of claim 26, wherein the thionin is synthesized.

33. The method of claim 28, wherein the second stain comprises reagent alcohol, eosin Y, thionin, and light or fast green.

34. The method of claim 26, wherein the thionin-phenol derivative solution binds preferentially to nuclear cytological material relative to cytoplasmic cytological material.

35. The method of claim 28, wherein the second stain binds preferentially to cytoplasmic cytology material relative to nuclear cytological material.

36. The method of claim 28, wherein at least one of the thionin-phenol derivative solution and the second stain discernibly stain the cytological material in a visible light range.

37. The method of claim 28, wherein both the thionin-phenol derivative solution and the second stain discernibly stain the cytological material in a visible light range.

38. The method of claim 26, further comprising the step of rinsing the cytological material after staining the cytological material.

39. The method of claim 28, wherein the second stain contains thionin to retard thionin depletion during the rinsing step.

40. The method of claim 28, wherein the cytological material is rinsed in an alcohol bath.

41. A method of imaging a specimen, the method comprising the steps of:

loading a specimen stained with a thionin-phenol solution into an optical instrument wherein the thionin-phenol solution comprises a phenol derivative;

illuminating the specimen; and

imaging the specimen.

42. The method of claim 41, further comprising the step of processing the image of the specimen.

43. A system for imaging a specimen stained with a thionin-phenol derivative solution, the system comprising:

an optical instrument;

at least one light source;

a processor; and

the stained specimen.