Radiant energy – Luminophor irradiation – With ultraviolet source
Patent
1993-04-12
1996-04-02
Willis, Davis L.
Radiant energy
Luminophor irradiation
With ultraviolet source
2504591, 356318, G01N 2164
Patent
active
055043372
DESCRIPTION:
BRIEF SUMMARY
BACKGROUND OF THE INVENTION
The present invention relates to a flow cytometry method and apparatus for distinguishing and/or characterizing cells or particles on the basis of measured fluorescence lifetimes. More particularly, the present invention concerns a flow cytometry method and apparatus for distinguishing and characterizing a particle or cell which has been labeled or associated with one or more fluorophores having lifetimes which are modified due to one or more characteristics or properties of a cell or particle with which the fluorophore is associated, wherein the lifetime measurments are independent of the intensity of the detected fluorescence emission.
Research involving the study and analysis of cells, generally known as cytology, employs a variety of analytical techniques for identifying and enumerating the subpopulations of cells in a specimen under study. For example, cytological materials may be examined to detect the presence of cancerous or malignant cells, or characteristics of the cells within a specimen. For purposes of analysis, the cells may be labeled with a variety of fluorescent materials, conventionally known as fluorophores or fluorescent probes, which have an identified affinity for cells or cell components which are of interest to an analysis. The fluorophores will emit a particular fluorescence radiation when stimulated by light at a wavelength corresponding to the excitation wavelength of the fluorophore. The wavelength and/or intensity of the emitted light has been used to analyze a subpopulation of cells, wherein different fluorophores can be used to distinguish subpopulations of fluorophore-labeled cells.
The study of collections of multiple cells using fluorescent spectroscopy has obvious limitations. For example, an accurate determination of the number of cells in a subpopulation having a given characteristic cannot be made and, more importantly, the subpopulations cannot be separated for further analysis. In order to permit the measurement and analysis of a population of cells (or any biological particle such as isolated nuclei, chromosome preparations or neurobiological organisms) on an individual basis, fluorescence flow cytometry has been employed.
Fluorescence flow cytometry, which involves the intensity and/or wavelength measurement of fluorescence emissions from individual cells labeled with a fluorophore while the cells are flowing in a liquid or gaseous medium past an observation point, permits analysis of individual cells as well as sorting of the cells based upon that analysis. A description of examples of fluorescence flow cytometry appears in "Practical Flow Cytometry", 2d. ed., H. Shapiro (1988), Liss & Co, the entire contents of which are herein incorporated by reference.
A conventional flow cytometer has a sample handling and delivery system which collects the cell population into a stream of individual cells which is directed one cell at a time past the observation point of the flow cytometer. When a liquid medium is used, the stream containing the samples is sheathed by a surrounding fluid stream to insure that only single cells pass the observation point.
A conventional flow cytometer also has a parameter detection system which can include a focused light source, typically a laser, that directs a narrow beam of light at a predetermined wavelength to the observation point where an individual fluorophore-labeled cell passing through the point may be illuminated, resulting in a fluorescence emission from the fluorophore label. The parameter detection system also includes collection optics and optical transducers, such as photomultipliers and detectors that receive the fluorescence emission at the observation point and convert it to electrical signals which are representative of the intensity and/or wavelength of the emitted light.
Where the cells have been tagged with a fluorophore having an affinity for a particular characteristic or composition of the cell, as well as being excited at the wavelength of light emitted by the laser, the light emission
REFERENCES:
patent: 5315122 (1994-05-01), Pinsky et al.
patent: 5317162 (1994-05-01), Pinsky et al.
Berndt Klaus W.
Hoffman Robert A.
Lakowicz Joseph R.
Pinsky Bertram G.
Hanig Richard
Lakowicz Joseph R.
Willis Davis L.
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