Methods for identifying stem cells by detecting fluorescence of cells and syncytia
AssigneeMassachusetts Institute of Technology
InventorsElena V. Gostjeva, William G. Thilly
Life of this patent
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Abstract
Methods are disclosed for the characterization of the stage of development or pathology of a tissue sample, and for identifying pluripotent metakaryotic stem cells, comprising detecting fluorescence of cells and syncytia in fixed samples treated with a non-fluorescent Schiff's base reagent in the absence of extraneous or exogenously added fluorescent dyes.
Claims28
A method for identifying a metakaryotic stem cell of interest having a bell-shaped nucleus, or identifying a multinuclear syncytium of interest, wherein the syncytium of interest comprises one or more metakaryotic stem cell nuclei having a bell-shape, the method comprising fixing a sample of cells and treating said sample with a non-fluorescent Schiff's base reagent in the absence of extraneous or exogenously added fluorescent dyes and visualizing fluorescent balloon-like cytoplasmic structures of a metakaryotic stem cell or fluorescence of the cytoplasm surrounding one or more stem cell nuclei within the syncytium, thereby identifying the metakaryotic stem cell of interest, or identifying a multinuclear syncytium of interest wherein the syncytium of interest comprises one or more bell-shaped stem cell nuclei.
A method for diagnosing preneoplasia or neoplasia in an adult mammal suspected of having preneoplasia or neoplasia, the method comprising fixing a sample of cells from the mammal and treating said sample with a non-fluorescent Schiff's base reagent in the absence of extraneous or exogenously added fluorescent dyes and detecting fluorescent balloon-like cytoplasmic structures of a metakaryotic stem cell or fluorescence of the cytoplasm surrounding one or more stem cell nuclei within a syncytium, wherein the stem cell nuclei are bell-shaped, whereby detecting one or more fluorescent metakaryotic stem cells having a bell-shaped nucleus or detecting one or more fluorescent syncytia comprising one or more stem cell nuclei having a bell-shape in the absence of extraneous or exogenously added fluorescent dyes is indicative of preneoplasia or neoplasia in the adult mammal.
A method for identifying a molecular marker specific to a metakaryotic stem cell or a syncytium comprising one or more stem cell nuclei, wherein the stem cell nucleus is bell-shaped, the method comprising comparing the molecular markers of a metakaryotic stem cell or the molecular markers of a syncytium comprising one or more stem cell nuclei with the molecular markers of a non-metakaryotic cell.
A method of identifying an agent which inhibits the proliferation of metakaryotic stem cells, comprising:
- a) treating in vitro a tissue or cell sample with a candidate agent, wherein the tissue or cell sample comprises a metakaryotic stem cell or a syncytium comprising one or more stem cell nuclei, wherein the stem cell nucleus is bell-shaped;
- b) fixing the treated tissue or cell sample and treating said sample with a non-fluorescent Schiff's base reagent in the absence of extraneous or exogenously added fluorescent dyes;
- c) detecting the presence, intensity, or number of fluorescent balloon-like cytoplasmic structures of a metakaryotic stem cell or fluorescence of the cytoplasm surrounding one or more stem cell nuclei within a syncytium;
- and
- d) comparing the presence, intensity, or number of fluorescent balloon-like cytoplasmic structures of a metakaryotic stem cell or fluorescence of the cytoplasm surrounding one or more stem cell nuclei within the syncytium in the sample treated with the candidate agent with a control,
- wherein a decrease in, or no increase in, the number of, or intensity of fluorescent stem cells, or syncytia containing one or more stem cell nuclei, is indicative that the agent inhibits the proliferation of metakaryotic stem cells.
A method of identifying an anti-tumorigenic agent, comprising:
- a) treating in vitro a tumor tissue or tumor cell sample with a candidate agent;
- b) fixing the treated tumor tissue or tumor cell sample and treating said sample with a non-fluorescent Schiff's base reagent in the absence of extraneous or exogenously added fluorescent dyes;
- c) detecting the presence, intensity, or number of fluorescent balloon-like cytoplasmic structures of a metakaryotic stem cell or fluorescence of the cytoplasm surrounding one or more stem cell nuclei within a syncytium comprising one or more stem cell nuclei, wherein the stem cell nucleus is bell-shaped;
- and
- d) comparing the presence, intensity, or number of fluorescent balloon-like cytoplasmic structures of a metakaryotic stem cell or fluorescence of the cytoplasm surrounding one or more stem cell nuclei within the syncytium in a sample treated with the candidate agent with a control,
- wherein a decrease in, or no increase in, the number of, or intensity of fluorescent stem cells, or syncytia comprising one or more stem cell nuclei, is indicative that the agent is an anti-tumorigenic agent.
A method of identifying an agent which inhibits the proliferation of stem cells, comprising:
- a) treating a test mammal with a candidate agent;
- b) fixing a sample of cells or tissue from the test mammal and treating said sample with a non-fluorescent Schiff's base reagent in the absence of extraneous or exogenously added fluorescent dyes;
- c) detecting the presence, intensity, or number of fluorescent balloon-like cytoplasmic structures of a metakaryotic stem cell or fluorescence of the cytoplasm surrounding one or more stem cell nuclei within a syncytium, wherein the stem cell nucleus is bell-shaped;
- and
- d) comparing the presence, intensity, or number of fluorescent balloon-like cytoplasmic structures of a metakaryotic stem cell or fluorescence of the cytoplasm surrounding one or more stem cell nuclei within the syncytium in the sample of cells or tissue from the test mammal treated with the candidate agent with a control,
- wherein a decrease in, or no increase in, the number of, or intensity of fluorescent stem cells, or syncytia containing one or more stem cell nuclei, is indicative that the agent inhibits the proliferation of metakaryotic stem cells.
A method of identifying an anti-tumorigenic agent comprising:
- a) treating a mammal having a tumor with a candidate agent;
- b) fixing a sample of tumor cells or tumor tissue from the mammal and treating said sample with a non-fluorescent Schiff's base reagent in the absence of extraneous or exogenously added fluorescent dyes;
- c) detecting the presence, intensity, or number of fluorescent balloon-like cytoplasmic structures of a metakaryotic stem cell or fluorescence of the cytoplasm surrounding one or more stem cell nuclei within a syncytium, wherein the stem cell nucleus is bell-shaped;
- and
- d) comparing the presence, intensity, or number of fluorescent balloon-like cytoplasmic structures of a metakaryotic stem cell or fluorescence of the cytoplasm surrounding one or more stem cell nuclei within the syncytium in the sample of tumor cells or tumor tissue from the mammal treated with the candidate agent with a control,
- wherein a decrease in, or no increase in, the number of, or intensity of fluorescent stem cells, or syncytia comprising one or more stem cell nuclei, is indicative that the agent is an anti-tumorigenic agent.
Dependencies are inferred from the claim text.