Method for detecting nucleosomes containing nucleotides

Abstract

The invention relates to a method for detecting and measuring the presence of mono-nucleosomes and oligo-nucleosomes and nucleosomes that contain particular nucleotides and the use of such measurements for the detection and diagnosis of disease. The invention also relates to a method of identifying nucleosome associated nucleotide biomarkers for the detection and diagnosis of disease and to biomarkers identified by said method.

It is claimed: 
     
         1 . A method for detecting cell-free nucleosomes with a 5-methylcytosine or a 5-hydroxymethylcytosine DNA base in a blood, serum or plasma sample which comprises cell-free nucleosomes, comprising:
 contacting the blood, serum or plasma sample with a first binding agent which binds to the cell free nucleosomes and a second binding agent which binds to a 5-methylcytosine or a 5-hydroxymethylcytosine DNA base; and   detecting cell-free nucleosomes with a 5-methylcytosine or 5-hydroxymethylcytosine DNA base that bind to the first binding agent and the second binding agent.   
     
     
         2 . The method of  claim 1 , wherein the first binding agent is an antibody. 
     
     
         3 . The method of  claim 2 , wherein the second binding agent is an antibody. 
     
     
         4 . The method of  claim 1 , wherein the second binding agent is an antibody. 
     
     
         5 . The method of  claim 1 , wherein the sample is serum. 
     
     
         6 . The method of  claim 1 , wherein the sample is plasma. 
     
     
         7 . A method to discriminate a blood, serum or plasma sample as from a diseased subject or a healthy subject, comprising:
 providing a blood, serum or plasma sample from the subject;   contacting the sample with a first binding agent which binds to cell free nucleosomes and with a second binding agent which binds to a 5-methylcytosine or a 5-hydroxymethylcytosine DNA base; and   detecting cell-free nucleosomes with a 5-methylcytosine or 5-hydroxymethylcytosine DNA base that bind to the first binding agent and the second binding agent to discriminate the sample as from a healthy subject or a diseased subject.   
     
     
         8 . The method of  claim 7 , wherein the first binding agent is an antibody. 
     
     
         9 . The method of  claim 8 , wherein the second binding agent is an antibody. 
     
     
         10 . The method of  claim 7 , wherein the second binding agent is an antibody. 
     
     
         11 . The method of  claim 7 , wherein the sample is serum. 
     
     
         12 . The method of  claim 7 , wherein the sample is plasma. 
     
     
         13 . The method of  claim 7 , wherein said detecting discriminates the sample as from a subject with a disease selected from the group consisting of cancer, cardiomyopathy, systemic lupus erythematosus, colitis, chronic obstructive pulmonary disorder, Crohn's disease and rheumatoid arthritis. 
     
     
         14 . A method for detecting or diagnosing a disease status in a subject, comprising:
 (i) detecting or measuring a level or type of cell free nucleosomes containing a particular DNA base, nucleotide or nucleoside in a body fluid of the subject;   (ii) comparing the detecting or measuring of step (i) to the level or type of cell free nucleosomes containing the particular DNA base, nucleotide or nucleoside in a body fluid of a healthy subject or a control subject; and   (iii) using the difference between the levels or types detected in diseased and control subjects to detect or identify disease status in the subject.   
     
     
         15 . A kit detecting cell-free nucleosomes with a 5-methylcytosine or a 5-hydroxymethylcytosine DNA base in a blood, serum or plasma sample which comprises cell-free nucleosomes, comprising:
 a first binding agent specific for the 5-methylcytosine or a 5-hydroxymethylcytosine DNA base,   a second binding agent which binds to cell free nucleosomes; and   instructions for use.