Apoptosis marker antibodies and methods of use

Abstract

Disclosed are antibodies that specifically recognize the new amino terminus of a protein cleaved by a protease during apoptosis. Methods of using and making the antibodies are also provided. The antibodies are particularly useful in methods of detecting apoptosis and testing candidate compounds for enhancing or inhibiting apoptosis.

1 . A method of diagnosing a disease, disorder, or condition associated with cell apoptosis, said method comprising: 
 (a) contacting a sample of cells or tissue from a patient with an antibody immunoreactive with a neoepitope produced in a cell undergoing apoptosis, said neoepitope comprising an amino terminus produced by cleavage of a protein by a protease, wherein said antibody is not immunoreactive with the protein when not cleaved by the protease; and    (b) detecting said antibody bound to the neoepitope, wherein detecting said antibody bound to the neoepitope is indicative of a disease, disorder, or condition associated with cell apoptosis.    
     
     
         2 . A method for screening compounds to identify an inhibitor of apoptosis, said method comprising: 
 (a) exposing a first and second sample of cells to conditions known to activate apoptosis in the cells;    (b) contacting the first sample with a test compound;    (c) contacting the first and second samples with an antibody immunoreactive with a neoepitope produced in a cell undergoing apoptosis, said neoepitope comprising an amino terminus produced by cleavage of a protein by a protease, wherein said antibody is not immunoreactive with the protein when not cleaved by the protease;    (d) detecting a level of said antibody bound to the neoepitope in said samples; and    (e) comparing the level of said first sample to said second sample, wherein said test compound inhibits apoptosis if the level of the antibody in the first sample is less than the level of the antibody in the second sample.    
     
     
         3 . A method for screening compounds to identify an inducer of apoptosis, said method comprising: 
 (a) contacting the first sample with a test compound;    (b) contacting the first and second samples with an antibody immunoreactive with a neoepitope produced in a cell undergoing apoptosis, said neoepitope comprising an amino terminus produced by cleavage of a protein by a protease, wherein said antibody is not immunoreactive with the protein when not cleaved by the protease;    (c) detecting a level of said antibody bound to the neoepitope in said samples; and    (d) comparing the level of said first sample to said second sample, wherein said test compound induces apoptosis if the level of the antibody in the first sample is greater than the level of the antibody in the second sample.    
     
     
         4 . A kit for detecting apoptosis-generated protein fragments in a sample, said kit comprising an antibody immunoreactive with a neoepitope produced in a cell undergoing apoptosis, said neoepitope comprising an amino terminus produced by cleavage of a protein by a protease, wherein said antibody is not immunoreactive with the protein when not cleaved by the protease.  
     
     
         5 . The kit of  claim 4 , wherein the antibody comprises a label.  
     
     
         6 . The kit of  claim 4  further comprising a labeled secondary antibody that is immunoreactive with the antibody.  
     
     
         7 . The kit of  claim 4  further comprising an antibody immunoreactive with a surface antigen.  
     
     
         8 . A kit for evaluating biological material, said kit comprising: 
 an antibody immunoreactive with a neoepitope in a biological material comprising a cell in which apoptosis has been initiated, said neoepitope comprising an amino terminus produced by cleavage of a protein by a protease, wherein said antibody is not immunoreactive with the protein when not cleaved by the protease.    
     
     
         9 . The kit of  claim 8 , wherein the cell is a mammalian cell.  
     
     
         10 . The kit of  claim 8 , wherein the biological material is tissue from a tumor.  
     
     
         11 . The kit of  claim 8 , further comprising an antibody immunoreactive with a proliferation marker.  
     
     
         12 . The kit of  claim 8 , further comprising reagents for the TdT-mediated dUTP Nick-End Labeling reaction.