Methods for Identifying Stem Cells Based on Nuclear Morphotypes

Abstract

Methods for identifying stem cells and other cells specific to embryogenesis and carcinogenesis, classifying tissue samples, diagnosing precancerous and cancerous or atherosclerotic lesions, testing the value of anticancer agents, discovering macromolecules specifically expressed in particular cell types, using stem cells in restorative tissue therapy as well as methods for preparing tissue samples so heteromorphic nuclear morphotypes remain intact are disclosed.

What is claimed is: 
     
         1 . A method for preparing a mammalian tissue sample suitable for the identification of cells comprising bell-shaped nuclei, comprising:
 a) disrupting cellular adhesions of cellular sheets of the tissue sample;   b) spreading the cells with disrupted adhesions onto a hard surface; and   c) artificially staining the cells and/or macromolecules of the sample, thereby allowing visualization of nuclei,   
       wherein the structural integrity of the nuclei of the cells remains intact, thus rendering the sample suitable for the identification of bell-shaped nuclei. 
     
     
         2 . The method of  claim 1 , wherein the tissue sample forms a layer on the microscope slide of about 0.5 millimeters. 
     
     
         3 . The method of  claim 1 , wherein the tissue sample forms a layer greater than about 50 microns. 
     
     
         4 . The method of  claim 1 , wherein the sample is a tissue sample obtained by surgical excision. 
     
     
         5 . The method of  claim 1 , wherein the sample is physically or chemically fixed prior to cellular degradation of nuclei. 
     
     
         6 . The method of  claim 5 , wherein the sample is frozen. 
     
     
         7 . The method of  claim 5 , wherein the sample is treated with one or more chemical fixing agents selected from the group consisting of: alcohols, aldehydes, organic acids and combinations thereof. 
     
     
         8 . The method of  claim 7 , wherein the fixing agent comprises methanol and acetic acid. 
     
     
         9 . The method of  claim 1 , wherein the cells of the sample are partially dissociated by tissue maceration and spreading. 
     
     
         10 . The method of  claim 1 , wherein DNA is stained, thereby allowing visualization of nuclei. 
     
     
         11 . The method of  claim 4 , wherein the tissue sample is fixed within 30 minutes of surgical removal. 
     
     
         12 . The method of  claim 1 , wherein the tissue sample is obtained from a mammal. 
     
     
         13 . The method of  claim 12 , wherein the mammal is selected from the group consisting of:
 primates, rodents, canines, felines, porcines, ovines, bovines and rabbits.   
     
     
         14 . The method of  claim 13 , wherein the mammal is a human. 
     
     
         15 . The method of  claim 1 , wherein the class or classes of nuclear morphotypes further comprise cigar-shaped, condensed spherical, spherical, oval, sausage-shaped, kidney-shaped, and bullet-shaped nuclei. 
     
     
         16 . The method of  claim 1 , wherein the nuclei are contained in multinuclear syncytia or in mononuclear cells. 
     
     
         17 . The method of  claim 1 , wherein the presence of non-spherical and non-oval nuclei in blood vessel wall tissue is indicative of an incipient atherosclerotic condition.