Method and apparatus for sorting cells
First Claim
1. A method of selecting a first set of cells from a population of cells including the first set of cells and a second set of cells, the method comprising:
- labeling the population of cells so that the first set of cells can be distinguished from the second set of cells;
providing a first flow path having a distal end, a proximal end, an interrogation area disposed between the proximal end and the distal end, and a flow axis;
creating a sheath flow of a sheath fluid through the first flow path, the sheath flow moving toward the proximal end at a first flow rate;
injecting into the sheath flow, at a point upstream from the proximal end, a specimen flow including the population of cells, the specimen flow having initially a second flow rate less than the first flow rate;
providing an excitation energy source, the energy emitted from the excitation energy source acting on individual cells as they pass through the interrogation area and causing emission or transmission of secondary radiation from the cells;
using an objective lens having an optical axis generally coaxially aligned with the flow axis to focus the secondary radiation from the individual cells as the cells pass through the interrogation area;
detecting the focused secondary radiation from the individual cells;
determining, from the detected secondary radiation, whether the individual cells are in the first set or the second set; and
selecting cells determined to be in the first set.
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Abstract
A method, apparatus, and system for a sorting flow cytometer include an objective lens having an optical axis coaxially aligned with the flow path at the focal point. A controllable energy source selectively alters an analyte according to a determination of whether the analyte is in a desired sub-population. In various embodiments, one or both of the emission from the controllable energy source and/or the emission from an illumination energy source passes through the objective lens. In some embodiments in which the emission from the controllable energy source passes through the objective lens, the objective lens may focus the emission from the controllable energy source at a different point than the focal point of a signal detected from the analyte and, in particular, at a point closer to the objective lens.
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Citations
43 Claims
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1. A method of selecting a first set of cells from a population of cells including the first set of cells and a second set of cells, the method comprising:
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labeling the population of cells so that the first set of cells can be distinguished from the second set of cells; providing a first flow path having a distal end, a proximal end, an interrogation area disposed between the proximal end and the distal end, and a flow axis; creating a sheath flow of a sheath fluid through the first flow path, the sheath flow moving toward the proximal end at a first flow rate; injecting into the sheath flow, at a point upstream from the proximal end, a specimen flow including the population of cells, the specimen flow having initially a second flow rate less than the first flow rate; providing an excitation energy source, the energy emitted from the excitation energy source acting on individual cells as they pass through the interrogation area and causing emission or transmission of secondary radiation from the cells; using an objective lens having an optical axis generally coaxially aligned with the flow axis to focus the secondary radiation from the individual cells as the cells pass through the interrogation area; detecting the focused secondary radiation from the individual cells; determining, from the detected secondary radiation, whether the individual cells are in the first set or the second set; and selecting cells determined to be in the first set. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11)
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12. An apparatus for detecting and selectively altering a desired sub-population of cells in a population of specimen cells, the apparatus comprising:
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a fluid flow path having; a first flow section having a flow axis, and a second flow section, the first and second flow sections intersecting at a measurement end of the first flow section; an interrogation area disposed at or near the measurement end of the first flow section; a sheath fluid input in fluid flow communication with the fluid flow path; a specimen input in fluid flow communication with the fluid flow path; an objective lens having a nominal focal point and an optical axis, and disposed at the measurement end of the first flow section, the objective lens aligned with the first flow section such that the nominal focal point is along the flow axis and in the interrogation area, and such that the optical axis is generally coaxially aligned with the flow axis; a detector disposed to detect light focused by the objective lens; a logic routine communicatively coupled to the detector, operable to determine whether a cell in the population of specimen cells is one of the desired sub-population of cells, and further operable to output a signal based on the determination of whether the cell is one of the desired sub-population of cells; and a controllable energy source communicatively coupled to the logic routine and operable to selectively alter either cells in the desired sub-population of cells or cells not in the desired sub-population of cells according to at least the signal output from the logic routine. - View Dependent Claims (13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23)
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24. A system for detecting and selectively altering a desired sub-population of cells in a population of specimen cells, the system comprising:
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fluid flow path having a flow axis; an interrogation area disposed within the fluid flow path; a sheath fluid input in fluid flow communication with the fluid flow path; a first pump in fluid flow communication with the sheath fluid input; a specimen fluid input in fluid flow communication with the fluid flow path; a second pump in fluid flow communication with the specimen fluid input; an objective lens having a nominal focal point and an optical axis, and disposed such that the nominal focal point is along the flow axis and in the interrogation area, and such that the optical axis is generally coaxially aligned with the flow axis in the interrogation area; a detector disposed to detect light focused by the objective lens; a controllable energy source; a processor communicatively coupled to a computer-readable storage medium, to the detector, and to the controllable energy source; and wherein the processor and the controllable energy source cooperate to selectively alter, according to an output from the processor, either cells in the desired sub-population of cells or cells not in the desired sub-population of cells. - View Dependent Claims (25, 26, 27, 28)
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29. A method of detecting and selectively altering a desired sub-population of cells in a population of specimen cells, the method embodied in a set of machine-readable instructions executed on a processor and stored on a tangible medium, the method comprising:
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controlling the flow of a population of specimen cells through a flow path having a flow axis; controlling an illumination source to illuminate an interrogation area through which the cells in the population of specimen cells pass; receiving data from a detector in an optical path having an objective lens, the objective lens having an optical axis and a nominal focal point, the optical axis generally coaxially aligned with the flow axis, the nominal focal point being within the interrogation area; determining from the received data the presence in the interrogation area of one of the cells in the population of specimen cells; determining from the received data whether the one of the specimen cells is one of the desired sub-population of cells; and controlling a cell selection energy source, according to at least the determination of whether the one of the specimen cells is part of the desired sub-population of cells. - View Dependent Claims (30, 31)
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32. A system for detecting and selectively altering a desired sub-population of cells in a population of specimen cells, the system comprising:
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a flow path having an interrogation area and a flow axis in the interrogation area; control means for controlling a flow of the population of specimen cells through the flow path; illumination means for illuminating the specimen cells as they pass through the interrogation area; an objective lens having an optical axis and a nominal focal point, the optical axis generally coaxially aligned with the flow axis, the nominal focal point being within the interrogation area; detection means for detecting energy focused by the objective lens and providing data related to the detected energy; processing means for receiving the data related to the detected energy and for determining whether individual specimen cells passing through the interrogation area are one of the desired sub-population; cell selection means for selectively irradiating the specimen cells; and cell selection control means for controlling the cell selection means according to at least the determination of whether the individual specimen cells are one of the desired sub-population. - View Dependent Claims (33, 34, 35)
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36. A process for detecting and selectively altering a desired sub-population of cells in a population of specimen cells, the process comprising:
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creating a flow carrying a generally single-file procession of specimen cells through a flow path; illuminating the specimen cells as the specimen cells pass through an interrogation area in the flow path; positioning an objective lens such that; an optical axis of the objective lens is generally coaxial with the flow path, the flow moves through the flow path toward the objective lens, and the objective lens has a nominal focal point in the interrogation area; detecting a parameter of individual specimen cells as the specimen cells pass through the interrogation area; interpreting the detected parameter of the individual specimen cells to determine whether the individual specimen cells are one of the desired sub-population; selectively derivatizing, killing, damaging, modifying, disrupting, or fragmenting one or more of the population of specimen cells according to the determination of whether the individual specimen cells are one of the desired sub-population of cells; and collecting the resulting population of processed cells. - View Dependent Claims (37, 38, 39, 40, 41, 42, 43)
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Specification