EP0121261A2
Expired / Lapsed
EP
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Method and apparatus for distinguishing subclasses of leukocytes 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 in a sample comprises passing particles in a sample, substantially one at a time, through an area of light detected at the particles. The volume of each particle is determined, preferably, as it passes through the area of focused light incident to the particles. Light scattered by each particle is detected as the particle passesthrough the area of light, over a wide angle relative to the direction of incident light. This method includes distinguishing different subpopulations of the particles by associating the determined volume of each particle with its detected light scatter. The simultaneous detection of particle volume, fluorescence and light scatter is determined in a single pass through flow cytometry equipment. An apparatus for carrying out the abovedescribed method is also within the purview of the present invention.
Inventors
- 👤 Bernard A. Shoor
Bibliographic Details
Application Number
—
Jurisdiction
EP
Data Source
google_patents
Last Synchronized
2026-10-27 19:41
CPC Classifications
G; G01; G01N; G01N15/00; G01N15/10; G01N15/14; G01N15/1456; G01N15/1459; G01N15/1031; G01N15/12; G01N2015/1019; G01N2015/1477; G01N21/00; G01N21/62; G01N21/63; G01N21/64
Claims Analysis
Total Claims: 35
Claim 1
Independent Claim
127 words
A method for distinguishing subclasses of leukocytes in a sample obtained from blood comprising: passing leukocyte cells in said sample, substantially one at a time, through an area of light provided by an incident beam of illumination directed at an axis substantially orthogonally to the direction of leukocyte flow; determining the volume of each cell as it passes through said area by sensing variations in an electrical impedance associated with the cells during flow through said area; detecting light scattered by each cell, as it passes through said area, over a wide angle relative to the axis of the incident beam of illumination; and distinguishing different subclasses of the leukocyte cells by associating the determined volume of each cell as a function of its detected light scatter.
Claim 2
Independent Claim
127 words
A method for distinguishing subclasses of leukocytes in a sample obtained from blood comprising: passing leukocyte cells in said sample, substantially one at a time, through an area of light provided by an incident beam of illumination directed at an axis substantially orthogonally to the direction of leukocyte flow; determining the volume of each cell as it passes through said area by sensing variations in an electrical impedance associated with the cells during flow through said area; detecting light scattered by each cell, as it passes through said area, over a wide angle relative to the axis of the incident beam of illumination; and distinguishing different subclasses of the leukocyte cells by associating the determined volume of each cell as a function of its detected light scatter.
Claim 3
Dependent Claim
34 words
The method of Claim 1 wherein said cells are passed through a transparent flow chamber having an orifice therethrough whereby the variations in electrical impedance are sensed when the cells flow through said orifice.
Claim 4
Dependent Claim
43 words
The method of Claim 2 wherein said cells are passed in a bi-component stream of liquid through said orifice, said stream having a particle free outer sheath liquid capable of carrying an electrical current and a coaxially arranged inner liquid containing said cells.
Claim 5
Dependent Claim
32 words
The method of Claim 1 wherein light scattered by each cell is detected at a wide angle ranging between 65° and 115° relative to the axis of the incident beam of illumination.
Claim 6
Dependent Claim
28 words
The method of Claim 4 wherein light scattered by each cell is detected on an axis substantially 90° relative to the axis of the incident beam of illumination.
Claim 7
Dependent Claim
19 words
The method of Claim 1 wherein said distinguishing step includes the distinguishing of lymphocytes from monocytes and granular leukocytes.
Claim 8
Dependent Claim
20 words
The method of Claim 1 wherein said passing step includes providing a beam of incoherent illumination to said flowing cells.
Claim 9
Dependent Claim
70 words
The method of Claim 1 which further includes: l abeling the leukocytes in said sample with a fluorochrome having a predetermined flourescence response to specific optical stimulation provided by said beam of illumination; detecting the fluorescence emitted by . the fluorochromes as said cells pass through said area of light; and distinguishing different subclasses of the leukocyte cells based on the fluorescence, volume and light scatter associated with each cell.
Claim 10
Dependent Claim
30 words
The method of Claim 8 wherein the fluorescence emitted by each cell is determined simultaneously with the determination of cell volume and the detection of light scattered by each cell.
Claim 11
Independent Claim
81 words
A method for distinguishing multiple subpopulations of particles in a sample comprising: passing particles in a sample, substantially one at a time, through an area of light directed at said particles; determining the volume of each particle; detecting light scattered by each particle, as it passes through said area, over a wide angle relative to said direction of light; and distinguishing different subpopulations of said particles by associating the determined volume of each particle with its detected wide angle light scatter.
Claim 12
Independent Claim
81 words
A method for distinguishing multiple subpopulations of particles in a sample comprising: passing particles in a sample, substantially one at a time, through an area of light directed at said particles; determining the volume of each particle; detecting light scattered by each particle, as it passes through said area, over a wide angle relative to said direction of light; and distinguishing different subpopulations of said particles by associating the determined volume of each particle with its detected wide angle light scatter.
Claim 13
Dependent Claim
18 words
The method of Claim 10 wherein the volume of each particle-is determined as it passes through said area.
Claim 14
Dependent Claim
47 words
The method of Claim 10 which further includes labeling said particles with a marking agent, sensing the marking agent on particles passing through said area, and distinguishing different subpopulations of particles by simultaneously determining volume, detecting light scatter and sensing the marking agent associated with each particle.
Claim 15
Dependent Claim
33 words
The method of Claim 12 wherein said labeling step includes labeling said particles with a plurality of different marking agents, and sensing the different marking agents to distinguish different subpopulations of said particles.
Claim 16
Independent Claim
130 words
A apparatus for distinguishing subclasses of leukocytes in a sample obtained from blood comprising: an electrically-sensitive orifice; means for moving leukocyte cells in said sample, substantially one at a time, in a flow path through said orifice to cause electrical signals to be generated relating to the volume of said cells; means for providing an incident beam of illumination on an axis substantially orthogonally to the direction of said cell flow through said orifice; means for detecting light scattered by each cell, as it passes through said area, over a wide angle relative to. the axis of the beam of illumination; and means for distinguishing different subclasses of leukocyte cells by associating the electrical signals relating to the volume of each cell as a function of its detected light scatter.
Claim 17
Independent Claim
130 words
A apparatus for distinguishing subclasses of leukocytes in a sample obtained from blood comprising: an electrically-sensitive orifice; means for moving leukocyte cells in said sample, substantially one at a time, in a flow path through said orifice to cause electrical signals to be generated relating to the volume of said cells; means for providing an incident beam of illumination on an axis substantially orthogonally to the direction of said cell flow through said orifice; means for detecting light scattered by each cell, as it passes through said area, over a wide angle relative to. the axis of the beam of illumination; and means for distinguishing different subclasses of leukocyte cells by associating the electrical signals relating to the volume of each cell as a function of its detected light scatter.
Claim 18
Dependent Claim
14 words
The apparatus of Claim 14 wherein said orifice is in a transparent flow chamber.
Claim 19
Dependent Claim
24 words
The apparatus of Claim 15 which includes a pair of electrodes, each positioned on opposite sides of said orifice for generating a potential thereacross.
Claim 20
Dependent Claim
20 words
The apparatus of Claim 14 wherein the means for providing said beam of illumination is a source of incoherent light.
Claim 21
Dependent Claim
23 words
The apparatus of Claim 17 wherein said providing means provides a light beam substantially conically configured as it is directed toward said orifice.
Claim 22
Dependent Claim
14 words
The apparatus of Claim 17 wherein said light source is a mercury arc lamp.
Claim 23
Dependent Claim
33 words
The apparatus of Claim 14 wherein said means for detecting scattered light detects such light at an angle ranging between 65° and 115° relative to the axis of the incident beam of illumination.
Claim 24
Dependent Claim
31 words
The apparatus of Claim 20 wherein said means for detecting scattered light detects such light on an axis substantially at 90° relative to the axis of the incident beam of illumination.
Claim 25
Dependent Claim
22 words
The apparatus of Claim 14 wherein said means for detecting includes a photomultiplier device capable of converting optical signals into electrical signals.
Claim 26
Dependent Claim
32 words
The apparatus of Claim 22 wherein said means for detecting further includes a light scatter collection lens positioned near said orifice for focusing light scattered by said cells toward said photomultiplier device.
Claim 27
Dependent Claim
33 words
The apparatus of Claim 23 wherein said means for detecting further includes an image aperture positioned in the image plane of said light scatter collection lens between said lens and said photomultiplier device.
Claim 28
Dependent Claim
20 words
The apparatus of Claim 14 which further includes means for detecting fluorescence emitted by excited fluorochromes associated with said cells.
Claim 29
Dependent Claim
27 words
The apparatus of Claim 25 wherein said means for distinguishing associates the fluorescence, volume and wide angle light scatter of each cell in a . simultaneous determination.
Claim 30
Dependent Claim
27 words
The apparatus of Claim 25 wherein said means for detecting fluorescence includes a photodetector device capable of detecting light in a specific region of the color spectrum.
Claim 31
Dependent Claim
28 words
The apparatus of Claim 14 wherein said means for distinguishing includes display means for indicating the detected volume and light scatter distribution of leukocyte cells in the sample.
Claim 32
Independent Claim
89 words
An apparatus for distinguishing multiple subpopulations of particles in a sample comprising: means for passing particles in a sample, substantially one at a time, through an area of light directed at said particles; means for determining the volume of each particle; means for detecting light scattered by each particle, as it passes through said area, over a wide angle relative to said direction of light; and means for distinguishing different subpopulations of said particles by associating the determined volume of each particle with its detected wide angle light scatter.
Claim 33
Independent Claim
89 words
An apparatus for distinguishing multiple subpopulations of particles in a sample comprising: means for passing particles in a sample, substantially one at a time, through an area of light directed at said particles; means for determining the volume of each particle; means for detecting light scattered by each particle, as it passes through said area, over a wide angle relative to said direction of light; and means for distinguishing different subpopulations of said particles by associating the determined volume of each particle with its detected wide angle light scatter.
Claim 34
Independent Claim
87 words
An apparatus for distinguishing multiple subpopulations of particles in a sample which particles have been labeled-with-a marking agent comprising: means for passing said labeled particles in a sample, substantially one at a time, through an area of light directed at said particles; means for simultaneously determining the presence of the marking agent on, volume of, and light scattered by, each particle as it passes through said area; and means for distinguishing different subpopulations of particles based on the association of at least two of the above-determined parameters.
Claim 35
Independent Claim
87 words
An apparatus for distinguishing multiple subpopulations of particles in a sample which particles have been labeled-with-a marking agent comprising: means for passing said labeled particles in a sample, substantially one at a time, through an area of light directed at said particles; means for simultaneously determining the presence of the marking agent on, volume of, and light scattered by, each particle as it passes through said area; and means for distinguishing different subpopulations of particles based on the association of at least two of the above-determined parameters.
References Cited (Backward Citations)
No citation records indexed.
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