METHOD OF MEASURING IMMUNE ACTIVATION
The invention is directed to methods for measuring immune activation by the level of clonotypes having the same unique regions and different isotype-determining regions. In one aspect, the method of the invention comprises forming a sequence-based clonotype profile from a sample containing B lymphocytes, wherein each clonotype of such profile comprises a unique region, such as a portion of a VDJ segment, and an isotype determining region, such as a portion of a C gene segment. Immune activation is indicated whenever the level of such clonotypes exceeds an upper bound of a reference range determined from multiple individual measurements or population measurements.
1 . A method of detecting immune activation in an individual, the method comprising the steps of:
obtaining a sample of nucleic acids from lymphocytes of an individual, the sample comprising recombined sequences each including at least a portion of a C gene segment of a B cell receptor;
generating an amplicon from the recombined sequences, each sequence of the amplicon including a portion of a C gene segment;
sequencing the amplicon to generate a profile of clonotypes each comprising at least a portion of a VDJ region of a B cell receptor and at least a portion of a C gene segment; and
determining a level of clonotypes in the profile which have VDJ regions that are identical and C gene segments that are different.
2 . The method of claim 1 wherein said C gene segment is from a nucleotide sequence encoding an IgH chain of said B cell receptor.
3 . The method of claim 2 wherein said profile of clonotypes comprises at least 10 4 clonotypes.
4 . The method of claim 2 further including correlating with immune activation in said individual said level of said clonotypes which have VDJ regions that are identical and C segments that are different.
5 . The method of claim 4 wherein said level is correlated with immune activation whenever said level exceeds an upper bound of a reference range.
6 . The method of claim 5 wherein said reference range is based on a population average.
7 . The method of claim 1 wherein said lymphocytes of said individual are obtained from peripheral blood of said individual.
8 . A method of detecting immune activation in an individual, the method comprising the steps of:
obtaining a sample of nucleic acids from lymphocytes of an individual, the sample comprising recombined sequences each including at least a portion of a C gene segment of a B cell receptor;
amplifying the recombined sequences in a polymerase chain reaction comprising primers specific for the C gene segments to form an amplicon;
sequencing the amplicon to generate a clonotype profile wherein each clonotype comprises at least a portion of a VDJ region of a B cell receptor and at least a portion of a C gene segment;
determining a level of clonotypes in the clonotype profile which have VDJ regions that are identical and C gene segments that are different; and
correlating such level with immune activation in the individual whenever such level exceeds an upper bound of a reference range.
9 . The method of claim 8 wherein said C gene segment is from a nucleotide sequence encoding an IgH chain of said B cell receptor.
10 . The method of claim 8 wherein said profile of clonotypes comprises at least 10 4 clonotypes.