US5585469A
Expired / Lapsed
US
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Dyeing agent having at least two dyes for staining a biological sample and staining method employing the dyeing agent
Assignee: Hitachi Ltd
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Filing: 1994-09-30
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Granted: 1996-12-17
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Expires: 2014-09-30
Abstract
Abstract Dyeing agents which are excellent in visual recognition of discernible or tangible components in a sample cause no coagulation of proteins, sugars or glycoproteins dissolved in the sample. An apparatus for image analysis of flow type stain particles uses the dyeing agents, and particles of discernible components suspended in a flowing sample can be detected so that images of the particles can be efficiently photographed and the discernible components can thus be analyzed by image processing of the thus-obtained image.
Inventors
- 👤 Yasuaki Kojima
- 👤 Ryohei Yabe
- 👤 Mitsuo Hattori
- 👤 Kazumitsu Kawase
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-11-09 13:20
CPC Classifications
G; G01; G01N; G01N33/00; G01N33/48; G01N33/50; G01N33/52; G01N1/00; G01N1/28; G01N1/30; G01N15/00; G01N15/10; G01N15/14; Y; Y10; Y10S; Y10S436/00; Y10S436/805; Y10T; Y10T436/00; Y10T436/10; Y10T436/107497; Y10T436/25
Claims Analysis
Total Claims: 27
Claim 1
Independent Claim
124 words
A dyeing agent for staining a biological sample, which comprises an azo dye and a xanthene dye suitable for supravital staining of cells and tissue, and a pH buffer, said azo and xanthene dyes being nonpolar molecular dyes which, upon blending, cause no agglutination or sedimentation of the dyes and which do not precipitate or agglutinate sugars, proteins or glycoproteins dissolved in said biological sample; wherein the blended azo and xanthene dyes are suitable for staining at least two objects, to different respective degrees in color hue or in dyeing strength depending on the respective objects; and wherein the blended azo and xanthene dyes are suitable for staining respective components of a single object to different degrees in color hue or in dyeing strength.
Claim 2
Dependent Claim
124 words
A dyeing agent for staining a biological sample, which comprises an azo dye and a xanthene dye suitable for supravital staining of cells and tissue, and a pH buffer, said azo and xanthene dyes being nonpolar molecular dyes which, upon blending, cause no agglutination or sedimentation of the dyes and which do not precipitate or agglutinate sugars, proteins or glycoproteins dissolved in said biological sample; wherein the blended azo and xanthene dyes are suitable for staining at least two objects, to different respective degrees in color hue or in dyeing strength depending on the respective objects; and wherein the blended azo and xanthene dyes are suitable for staining respective components of a single object to different degrees in color hue or in dyeing strength.
Claim 3
Dependent Claim
23 words
A dyeing agent according to claim 1, wherein said azo dye is selected from the group consisting of Evans Blue and Trypan Blue.
Claim 4
Dependent Claim
22 words
A dyeing agent according to claim 1, wherein said xanthene dye is selected from the group consisting of Erythrosine, Phloxine and Eosin.
Claim 5
Dependent Claim
22 words
A dyeing agent according to claim 2, wherein said xanthene dye is selected from the group consisting of Erythrosine, Phloxine and Eosin.
Claim 6
Dependent Claim
28 words
A dyeing agent according to claim 4, wherein said pH buffer is selected from the group consisting of a phosphate buffer, a succinate buffer and a tris-acid buffer.
Claim 7
Dependent Claim
16 words
A dyeing agent according to claim 5, further comprising an antibacterial agent as a fixative agent.
Claim 8
Dependent Claim
27 words
A dyeing agent according to claim 6, wherein said antibacterial agent is selected from the group consisting of sodium azide, para-hydroxyphenylacetic acid, dehydroacetic acid and ethylenediaminetetraacetic acid.
Claim 9
Dependent Claim
71 words
A dyeing agent obtained by mixing one volume of about 0.2 to 10.0×10 -2 mol/l of Evans Blue or Trypan Blue with about 0.5 to 2.0 volumes of about 0.2 to 10.0×10 -2 mol/l of Erythrosine; and adjusting a pH of the system to 5.7 to 7.9 using 1/30 to 1/5 mols/l of a solvent selected from the group consisting of a phosphate buffer, a succinate buffer and a tris-acid buffer.
Claim 10
Dependent Claim
71 words
A dyeing agent obtained by mixing one volume of about 0.2 to 10.0×10 -2 mol/l of Evans Blue or Trypan Blue with about 0.5 to 2.0 volumes of about 0.2 to 10.0×10 -2 mol/l of Erythrosine; and adjusting a pH of the system to 5.7 to 7.9 using 1/30 to 1/5 mols/l of a solvent selected from the group consisting of a phosphate buffer, a succinate buffer and a tris-acid buffer.
Claim 11
Dependent Claim
12 words
A dyeing agent according to claim 1, further comprising a fixative agent.
Claim 12
Dependent Claim
111 words
A dyeing agent according to claim 9 obtained by mixing one volume of about 0.2 to 10.0×10 -2 mol/l of Evans Blue or Trypan Blue as the azo dye with about 0.5 to 2.0 volumes of about 0.2 to 10.0×10 -2 mol/l of Phloxine as the xanthene dye; adjusting a pH of the system to 5.7 to 7.9 using 1/30 to 1/5 mols/l of a solvent selected from the group consisting of a phosphate buffer, a succinate buffer and a tris-acid buffer; and further adding an agent selected from the group consisting of glutaraldehyde, formaldehyde and paraformaldehyde as the fixative agent to the mixture in a concentration of 0.02 to 5.0%.
Claim 13
Dependent Claim
13 words
A dyeing agent according to claim 9, further comprising a surface active agent.
Claim 14
Dependent Claim
129 words
A dyeing agent according to claim 11, which is obtained by mixing one volume of about 0.2 to 10.0×10 -2 mol/l of Evans Blue or Trypan Blue as the azo dye with about 0.5 to 2.0 volumes of about 0.2 to 10.0×10 -2 mol/l of Eosine as the xanthene dye; adjusting a pH of the system to 5.7 to 7.9 using 1/30 to 1/5 mols/l of a solvent selected from the group consisting of a phosphate buffer, a succinate buffer and a tris-acid buffer; and further adding an agent selected from the group consisting of glutaraldehyde, formaldehyde and paraformaldehyde as the fixative agent to the mixture in a concentration of 0.02 to 5.0%, and sodium dodecyl sulfate as the surface active agent in a concentration of 0.01 to 0.5%.
Claim 15
Dependent Claim
21 words
A method for staining discernible urinary sediments in urine, comprising the step of using a dyeing agent according to claim 7.
Claim 16
Dependent Claim
43 words
A dyeing agent according to claim 8, wherein the dyeing agent is further obtained by mixing with the mixture one member selected from the group consisting of sodium azide, parahydroxyphenylacetic acid, dehydroacetic acid and ethylenediaminetetraacetic acid in a concentration of 0.01 to 1.0%.
Claim 17
Dependent Claim
43 words
A dyeing agent according to claim 10, wherein the dyeing agent is further obtained by mixing with the mixture one member selected from the group consisting of sodium azide, para-hydroxyphenylacetic acid, dehydroacetic acid and ethylenediaminetetraacetic acid in a concentration of 0.01 to 1.0%.
Claim 18
Dependent Claim
43 words
A dyeing agent according to claim 12, wherein the dyeing agent is further obtained by mixing with the mixture one member selected from the group consisting of sodium azide, para-hydroxyphenylacetic acid, dehydroacetic acid and ethylenediaminetetraacetic acid in a concentration of 0.01 to 1.0%.
Claim 19
Dependent Claim
109 words
A dyeing agent for staining a biological sample, obtained by mixing one volume of about 0.2 to 10.0×10 -2 mol/l of Evans Blue or Trypan Blue as a first dye with about 0.5 to 2.0 volumes of about 0.2 to 10.0×10 -2 mol/l of Phloxine as a second dye; adjusting a pH of the system to 5.7 to 7.9 using 1/30 to 1/5 mols/l of a solvent selected from the group consisting of a phosphate buffer, a succinate buffer and a tris-acid buffer; and further adding a fixative agent selected from the group consisting of glutaraldehyde, formaldehyde and paraformaldehyde to the mixture in a concentration of 0.02 to 5.0%.
Claim 20
Dependent Claim
109 words
A dyeing agent for staining a biological sample, obtained by mixing one volume of about 0.2 to 10.0×10 -2 mol/l of Evans Blue or Trypan Blue as a first dye with about 0.5 to 2.0 volumes of about 0.2 to 10.0×10 -2 mol/l of Phloxine as a second dye; adjusting a pH of the system to 5.7 to 7.9 using 1/30 to 1/5 mols/l of a solvent selected from the group consisting of a phosphate buffer, a succinate buffer and a tris-acid buffer; and further adding a fixative agent selected from the group consisting of glutaraldehyde, formaldehyde and paraformaldehyde to the mixture in a concentration of 0.02 to 5.0%.
Claim 21
Dependent Claim
13 words
A dyeing agent according to claim 17, further comprising a surface active agent.
Claim 22
Dependent Claim
105 words
A dyeing agent according to claim 17, wherein upon mixing the first and second dyes, the first and second dyes cause no agglutination or sedimentation of the first and second dyes, and do not precipitate or agglutinate sugars, proteins or glycoproteins dissolved in said biological sample; wherein the mixed first and second dyes are suitable for staining at least two objects to different respective degrees in color hue or in dyeing strength depending on the respective objects; and wherein the mixed first and second dyes are suitable for staining respective components of a single object to different degrees in color hue or in dyeing strength.
Claim 23
Dependent Claim
43 words
A dyeing agent according to claim 17, wherein the dyeing agent is further obtained by mixing with the mixture one member selected from the group consisting of sodium azide, para-hydroxyphenylacetic acid, dehydroacetic acid and ethylenediaminetetraacetic acid in a concentration of 0.01 to 1.0%.
Claim 24
Dependent Claim
125 words
A dyeing agent for staining a biological sample, obtained by mixing one volume of about 0.2 to 10.0×10 -2 mol/l of Evans Blue or Trypan Blue as a first dye with about 0.5 to 2.0 volumes of about 0.2 to 10.0×10 -2 mol/l of Eosine as a second dye; adjusting a pH of the system to 5.7 to 7.9 using 1/30 to 1/5 mols/l of a solvent selected from the group consisting of a phosphate buffer, a succinate buffer and a tris-acid buffer; and further adding a fixative agent selected from the group consisting of glutaraldehyde, formaldehyde and paraformaldehyde to the mixture in a concentration of 0.02 to 5.0%, and sodium dodecyl sulfate as a surface active agent in a concentration of 0.01 to 0.5%.
Claim 25
Dependent Claim
125 words
A dyeing agent for staining a biological sample, obtained by mixing one volume of about 0.2 to 10.0×10 -2 mol/l of Evans Blue or Trypan Blue as a first dye with about 0.5 to 2.0 volumes of about 0.2 to 10.0×10 -2 mol/l of Eosine as a second dye; adjusting a pH of the system to 5.7 to 7.9 using 1/30 to 1/5 mols/l of a solvent selected from the group consisting of a phosphate buffer, a succinate buffer and a tris-acid buffer; and further adding a fixative agent selected from the group consisting of glutaraldehyde, formaldehyde and paraformaldehyde to the mixture in a concentration of 0.02 to 5.0%, and sodium dodecyl sulfate as a surface active agent in a concentration of 0.01 to 0.5%.
Claim 26
Dependent Claim
105 words
A dyeing agent according to claim 21, wherein upon mixing the first and second dyes, the first and second dyes cause no agglutination or sedimentation of the first and second dyes and do not precipitate or agglutinate sugars, proteins or glycoproteins dissolved in said biological sample; wherein the mixed first and second dyes are suitable for staining at least two objects to different respective degrees in color hue or in dyeing strength depending on the respective objects; and wherein the mixed first and second dyes are suitable for staining respective components of a single object to different degrees in color hue or in dyeing strength.
Claim 27
Dependent Claim
43 words
A dyeing agent according to claim 21, wherein the dyeing agent is further obtained by mixing with the mixture one member selected from the group consisting of sodium azide, para-hydroxyphenylacetic acid, dehydroacetic acid and ethylenediaminetetraacetic acid in a concentration of 0.01 to 1.0%.
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USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
|---|---|---|---|---|---|
| 3.5 Years (Year 4) | 1999-12-17 | 2000-06-17 | 2000-12-17 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 2003-12-17 | 2004-06-17 | 2004-12-17 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 2007-12-17 | 2008-06-17 | 2008-12-17 | $7700.00 | Window Closed / Lapsed |