Rapid, multifluorescent TCRG Vγ and jγ typing: application to T cell acute lymphoblastic leukemia and to the detection of minor clonal populations

Detection of clonal T cell receptor γ (TCRG) gene rearrangements by PCR is widely used in both the diagnostic assessment of lymphoproliferative disorders and the follow-up of acute lymphoblastic leukaemia (ALL), when residual positivity in excess of 10−3 at morphological complete remission is increasingly recognised to be an independent marker of poor prognosis. This is largely based on specific detection of V–J rearrangements from childhood cases. We describe rapid, multifluorescent Vγ and Jγ PCR typing of multiplex amplified diagnostic samples, as applied to 46 T-ALL. These strategies allow selected analysis of appropriate cases, immediate identification of Vγ and Jγ segments in over 95% of alleles, improved resolution and precision sizing and a sensitivity of detection at the 10−2–10−3 level. We demonstrate preferential V–J combinations but no difference in V–J usage between children and adults, nor between SIL-TALI-negative and -positive cases. A combination of fluorescent multiplex and Vγ–Jγ-specific monoplex follow-up, as described here, will allow detection of both significant clonal evolution and of the diagnostic clone at a level of prognostic significance, by techniques which can readily be applied to large-scale prospective studies for which real-time analysis is required.