EP0121262A2
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EP
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Method and apparatus for distinguishing multiple subpopulations of cells in a sample
Assignee: Becton Dickinson and Co
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Filing: 1984-03-31
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Granted: —
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Expires: 2004-03-31
Abstract
Abstract A method for distinguishing multiple subpopulations of particles, such as cells, in a sample comprises selectively labeling particles in a sample with a plurality of different surface marking agents. Each marking agent has distinguishing, quantifiable marking characteristics. This method includes passing the labeled particles, substantially one at a time, successively through a plurality of areas of detection to detect the characteristics of one or more different marking agents on particles in each area of detection. A greater number of marking agents are detected than the number of detection areas through which particles pass. Differences among the particles are distinguished relative to the detected, different quantifiable characteristics of the marking agents. An apparatus for carrying out the above-described method is also within the purview of the present invention.
Inventors
- 👤 Michael R. Loken
- 👤 David R. Parks
- 👤 Richard R. Hardy
Bibliographic Details
Application Number
—
Jurisdiction
EP
Data Source
google_patents
Last Synchronized
2026-10-28 17:56
CPC Classifications
G; G01; G01N; G01N21/00; G01N21/62; G01N21/63; G01N21/64; G01N21/645; G01N15/00; G01N15/10; G01N15/14; G01N2021/6417; G01N2021/6419; G01N2021/6421; G01N21/6428; G01N2021/6439; G01N2021/6441; G01N33/00; G01N33/48; G01N33/50; G01N33/53; G01N33/531; G01N33/532; G01N33/533
Claims Analysis
Total Claims: 24
Claim 1
Independent Claim
136 words
A method for distinguishing multiple subpopulations of cells in a sample comprising: selectively labeling cells in a sample with at least three different immunofluorescent stains, each stain having predetermined, substantially separated emission spectra; passing said labeled cells, substantially one at a time, through a first area of focused optical stimulation to provide excitation energy to excite one or more of said different stains; passing said labeled cells, substantially one at a time, through at least one other area of focused optical stimulation to provide excitation energy to excite, in each other area, one or more of said different stains so-that-at least three different stains are excited by passing through said first and other areas; detecting the fluorescence emitted by the different excited stains; and distinguishing subpopulations of said cells relative to the detected fluorescence characteristics thereof.
Claim 2
Independent Claim
136 words
A method for distinguishing multiple subpopulations of cells in a sample comprising: selectively labeling cells in a sample with at least three different immunofluorescent stains, each stain having predetermined, substantially separated emission spectra; passing said labeled cells, substantially one at a time, through a first area of focused optical stimulation to provide excitation energy to excite one or more of said different stains; passing said labeled cells, substantially one at a time, through at least one other area of focused optical stimulation to provide excitation energy to excite, in each other area, one or more of said different stains so-that-at least three different stains are excited by passing through said first and other areas; detecting the fluorescence emitted by the different excited stains; and distinguishing subpopulations of said cells relative to the detected fluorescence characteristics thereof.
Claim 3
Dependent Claim
23 words
The method of Claim 1 wherein said cells are labeled with three different stains and there are two areas of focused optical stimulation.
Claim 4
Dependent Claim
19 words
The method of Claim 2 wherein the three different fluorochromes of the stains are fluorescein, phycoerythrin and Texas Red.
Claim 5
Dependent Claim
20 words
The method of Claim 2 wherein the three different fluorochromes of the stains are fluores-- cein, phycoerythrin and allo phycocyanin.
Claim 6
Dependent Claim
18 words
The method of Claim 2 wherein the three different fluorochromes of the stains are fluorescein, phycoerythrin and R-phycocyanin.
Claim 7
Dependent Claim
23 words
The method of Claim 1 wherein said cells are labeled with four different stains and there are two areas of focused optical stimulation.
Claim 8
Dependent Claim
21 words
The method of Claim 6 wherein the four different fluorochromes of the stains are fluorescein, phycoerythrin., Texas Red and allo phycocyanin.
Claim 9
Dependent Claim
20 words
The method of Claim 6 wherein the four different fluorochromes of the stains are fluorescein, phycoerythrin, R-phycocyanin and allo phycocyanin.
Claim 10
Dependent Claim
30 words
The method of Claim 1 wherein the focused optical stimulation in each of said areas is caused by providing coherent light at a given wavelength to each of said areas.
Claim 11
Dependent Claim
50 words
The method of Claim 9 wherein the coherent light provided to an area of stimulation is at a wavelength sufficiently separated from the wavelength of an adjacent spectral region of stimulation to be substantially outside of the spectral range of excitation and emission of the stains in an adjacent region.
Claim 12
Independent Claim
104 words
A method for distinguishing multiple subpopulations of particles in a sample comprising: selectively labeling particles in a sample with a plurality of different surface marking agents, each marking agent having distinguishing, quantifiable marking characteristics; passing said labeled particles, substantially one at a time, successively through a plurality of areas of detection to detect the characteristics of one or more different marking agents on particles in each area of detection to detect a greater number of marking agents than the number of said areas through which particles pass; and distinguishing differences among said particles relative to the detected, different, quantifi-able characteristics of said marking agents.
Claim 13
Independent Claim
104 words
A method for distinguishing multiple subpopulations of particles in a sample comprising: selectively labeling particles in a sample with a plurality of different surface marking agents, each marking agent having distinguishing, quantifiable marking characteristics; passing said labeled particles, substantially one at a time, successively through a plurality of areas of detection to detect the characteristics of one or more different marking agents on particles in each area of detection to detect a greater number of marking agents than the number of said areas through which particles pass; and distinguishing differences among said particles relative to the detected, different, quantifi-able characteristics of said marking agents.
Claim 14
Independent Claim
114 words
An apparatus for distinguishing multiple subpopulations of cells which have been selectively labeled with different immunofluorescent stains each having predetermined, substantially separated emission spectra comprising; means for moving said labeled cells, substantially one at a time, in a flow path; first means for exciting one or more immunofluorescent stain in a first area of optical stimulation along said flow path; second means for exciting one or more of said different immunofluorescent stains in a second area of optical stimulation along said flow path; means for detecting the fluorescence emitted by the different excited immunofluorescent stains associated with the cells; and means for distinguishing subpopulations of said cells relative to the detected fluorescence characteristics thereof.
Claim 15
Independent Claim
114 words
An apparatus for distinguishing multiple subpopulations of cells which have been selectively labeled with different immunofluorescent stains each having predetermined, substantially separated emission spectra comprising; means for moving said labeled cells, substantially one at a time, in a flow path; first means for exciting one or more immunofluorescent stain in a first area of optical stimulation along said flow path; second means for exciting one or more of said different immunofluorescent stains in a second area of optical stimulation along said flow path; means for detecting the fluorescence emitted by the different excited immunofluorescent stains associated with the cells; and means for distinguishing subpopulations of said cells relative to the detected fluorescence characteristics thereof.
Claim 16
Dependent Claim
29 words
The apparatus of Claim 12 wherein the first means and the second means for exciting each includes a source for providing coherent light to said respective areas of stimulation.
Claim 17
Dependent Claim
46 words
The apparatus of Claim 13 wherein the sources of coherent light are selected to provide light at wavelengths sufficiently separated from each other to be substantially outside of the spectral range of excitation and emission of the stains in each of -the respective areas of stimulation.
Claim 18
Dependent Claim
30 words
The apparatus of Claim 14 wherein, the first source of coherent light is an argon ion laser and the second source of coherent light is a rhodamine 6-G dye laser.
Claim 19
Dependent Claim
29 words
The apparatus of Claim 14 wherein the first source of coherent light is an argon ion laser and the second source of coherent light is a helium- neon laser.
Claim 20
Dependent Claim
41 words
The apparatus of Claim 12 wherein the means for detecting includes up to two photodetector devices associated with each of the first and second means for exciting, each such photodetector device capable of detecting light in a specifically defined color region.
Claim 21
Dependent Claim
20 words
The apparatus of Claim 12 wherein the means for distinguishing includes display means for indicating the detected subpopulations of cells.
Claim 22
Dependent Claim
20 words
The apparatus of Claim 12 which further includes means to determine the approximate number of cells detected in each subpopulation.
Claim 23
Independent Claim
111 words
An apparatus for distinguishing multiple subpopulations of particles in a sample which particles have been selectively labeled with different surface marking agents having distinguishable, quantifiable characteristics capable of being stimulated comprising: means for moving said labeled particles, substantially one at a time,-in a flow path; a plurality of .means for- stimulating: said agents on the particles moving in said flow path, each stimulating means being capable of stimulating the characteristics of more than one of the different marking agents on labeled particles; means for detecting the characteristics exhibited by said stimulated marking agents; and means for distinguishing differences among said particles relative to_the detected, different quantifiable characteristics of said marking agents.
Claim 24
Independent Claim
111 words
An apparatus for distinguishing multiple subpopulations of particles in a sample which particles have been selectively labeled with different surface marking agents having distinguishable, quantifiable characteristics capable of being stimulated comprising: means for moving said labeled particles, substantially one at a time,-in a flow path; a plurality of .means for- stimulating: said agents on the particles moving in said flow path, each stimulating means being capable of stimulating the characteristics of more than one of the different marking agents on labeled particles; means for detecting the characteristics exhibited by said stimulated marking agents; and means for distinguishing differences among said particles relative to_the detected, different quantifiable characteristics of said marking agents.
References Cited (Backward Citations)
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