Patent ID: 6087096
Filing Date: 2000-07-11
Classification: C12Q

Abstract:
A method of assaying for a T cell immunoproliferative condition in a human individual comprising the steps of:(a) obtaining cells from said individual, said cells including T lymphocytes (T cells);(b) generating an assay intrafamily gene fragment length profile from said cells for T cell receptor (TCR) third-complementarity-determining regions (CDR3) of an .alpha.-chain variable region gene family (V.alpha.) by:(i) isolating RNA from said cells;(ii) synthesizing cDNA from said RNA;(iii) performing a first polymerase chain reaction (PCR) with a reaction mixture that includes cDNA from step (ii) as a template; a family-specific TCR .alpha.-chain variable region oligonucleotide primer; and a TCR .alpha.-chain constant region oligonucleotide primer; wherein said first PCR amplifies DNA encoding TCR-CDR3 of an .alpha..beta. T cell receptor .alpha.-chain variable region gene family;(iv) performing a second PCR with a reaction mixture that includes amplified DNA from step (iii) as a template; the family-specific TCR .alpha.-chain variable region oligonucleotide primer; and a nested TCR .alpha.-chain constant region oligonucleotide primer; wherein said second polymerase chain reaction provides amplified DNA fragments encoding TCR-CDR3 of said .alpha..beta. T cell receptor .alpha.-chain variable region gene family;(v) separating DNA fragments from the second polymerase chain reaction by length; and(vi) determining a prevalence of DNA fragments separated according to step (v), to provide an intrafamily gene fragment length profile for said .alpha..beta. T cell receptor .alpha.-chain variable region gene family; and(c) comparing the assay profile of step (b) to a control intrafamily gene fragment length profile derived from blood cells of a healthy human subject, to determine a presence or an absence of at least one gene fragment length that is more prevalent in the assay profile than the control profile, wherein said control profile is for the same TCR-CDR3 .alpha.-chain variable region gene family as the assay profile, and wherein the presence of at least one gene fragment length that is more prevalent in the assay profile than the control profile is correlated to a T cell immunoproliferative condition.