Methods for Identifying Stem Cells by Detecting Fluorescence of Cells and Syncytia

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.

1 .- 12 . (canceled) 
     
     
         13 . 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.   
     
     
         14 . The method of  claim 13 , wherein fixing the treated tissue or cell sample comprises treating the sample of cells with Carnoy's fixative and reacting with Feulgen stain. 
     
     
         15 . The method of  claim 13 , wherein the control is an in vitro tissue or cell sample treated with a placebo, 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. 
     
     
         16 . 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.   
     
     
         17 . The method of  claim 16 , wherein fixing the treated tumor tissue or tumor cell sample comprises treating the sample of tumor cells with Carnoy's fixative and reacting with Feulgen stain. 
     
     
         18 . The method of  claim 16 , wherein the control is an in vitro tumor tissue or tumor cell sample treated with a placebo. 
     
     
         19 . A method of identifying an agent which inhibits the proliferation of metakaryotic 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.   
     
     
         20 . The method of  claim 19 , wherein fixing the treated tumor tissue or tumor cell sample comprises treating the sample of tumor cells with Carnoy's fixative and reacting with Feulgen stain. 
     
     
         21 . The method of  claim 19 , wherein the control is a sample of cells or tissue taken from the test mammal before treatment with the test agent, or the control is a sample of cells or tissue from a test mammal treated with a placebo. 
     
     
         22 . 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.   
     
     
         23 . The method of  claim 22 , wherein fixing the treated tumor tissue or tumor cell sample comprises treating the sample of tumor cells with Carnoy's fixative and reacting with Feulgen stain. 
     
     
         24 . The method of  claim 22 , wherein the control is a sample of tumor cells or tumor tissue taken from the mammal before treatment with the candidate agent, or the control is a sample of cells or tissue from a mammal treated with a placebo. 
     
     
         25 .- 28 . (canceled)