Search for cancer markers by a novel screening method

Abstract

The present invention relates to a method of concentrating low molecular weight peptides in the supernatant of serum-free cultured cells, said method comprising allowing the peptides to bind to a strong cation exchanger under an acid condition, and eluting them under an alkali condition to concentrate the peptide. Furthermore, peptides having the amino acid sequence as set forth in SEQ ID NO: 1 or 2, and a method of screening cancer markers using antibody to these peptides, are disclosed.

1 . A method of concentrating low molecular weight peptides in the supernatant of serum-free cultured cells, said method comprising: 
 a) allowing the peptides to bind to a strong cation exchanger under an acid condition, and    b) eluting them under an alkali condition to concentrate the peptides.    
     
     
         2 . A method of screening peptides that are specifically secreted from a certain cultured cell line, said method comprising concentrating peptides in two or more culture supernatants by the method according to  claim 1 , and then comparatively analyzing them using a mass spectrometer.  
     
     
         3 . A secretory peptide comprising part or all of an amino acid sequence that is substantially identical to the amino acid sequence as set forth in SEQ ID NO: 1.  
     
     
         4 . A secretory peptide comprising part or all of the amino acid sequence as set forth in SEQ ID NO: 1.  
     
     
         5 . A secretory peptide comprising part or all of an amino acid sequence that is substantially identical to the amino acid sequence as set forth in SEQ ID NO: 2.  
     
     
         6 . A secretory peptide comprising part or all of the amino acid sequence as set forth in SEQ ID NO: 2.  
     
     
         7 . A method of detecting or quantitating the peptide according to any of claims  3 - 6 .  
     
     
         8 . A diagnostic kit or a diagnostic reagent for diagnosing pancreatic cancer by detecting or quantitating the peptide according to any of claims  3 - 6 .  
     
     
         9 . A probe that binds to the peptide according to any of claims  3 - 6 .  
     
     
         10 . An antibody that has a binding affinity for the peptide according to any of claims  3 - 6 .  
     
     
         11 . The antibody according to  claim 10  which is a polyclonal antibody or a monoclonal antibody.  
     
     
         12 . A hybridoma that produces the monoclonal antibody according to  claim 11 .  
     
     
         13 . The method according to  claim 7  comprising the probe according to  claim 9  or the antibody according to  claim 10  or  11 .  
     
     
         14 . A diagnostic kit or a diagnostic reagent comprising the probe according to  claim 9  or the antibody according to  claim 10  or  11 .