Detection of T cell receptor beta (TCRB) gene rearrangement patterns in T cell malignancies by Southern blot analysis

Reliable detection of clonal T cell receptor beta (TCRB) gene rearrangement s is essential for clonality assessment of suspect T cell proliferations. S ince no appropriate Southern blot probes were available, we developed a new set of optimized TCRB gene probes. The TCRBJ1 and TCRBJ2 probes are positi oned just downstream of the J beta 1 and J beta 2 gene segments, respective ly, and can be used for detection of both incomplete D beta-J beta and comp lete V beta-J beta rearrangements, whereas the TCRBD1U and TCRBD2U probes u pstream of the D beta segments can be used to confirm incomplete D beta-J b eta rearrangements. Less frequently occurring V beta-D beta rearrangements can easily be detected via the downstream TCRBD1 and TCRBD2 probes. Althoug h both EcoRI and HindIII are appropriate restriction enzymes to be used in combination with all these probes, false-positivity due to partial digestio n of the EcoRI site in the J beta 2-C beta intron has to be excluded via th e TCRBC probe. Application of the TCRB probes in a large series of nearly 2 00 T cell malignancies revealed clonal rearrangements in all immature and m ature ICR alpha beta(+) T cell malignancies, in the vast majority of TCR ga mma delta(+) T cell acute lymphoblastic leukemias (T-ALL) (approximately 95 %) and even in most CD3(-) T-ALL (approximately 80%). TCRB gene rearrangeme nt patterns differed between the various categories of T cell malignancies. An increased frequency of complete V beta-J beta 1 rearrangements was obse rved in TCR alpha beta(+) T-ALL as compared to CD3- and TCR gamma delta(+) T-ALL (33% vs 16% and 11%, respectively), and also incompletely rearranged V-D, D-D, or D-J alleles in the beta 2 region occurred more frequently in T CR alpha beta(+) cases than in CD3- and TCR gamma delta(+) T-ALL (27% vs 15 % and 18%, respectively). Furthermore, in comparison to TCR alpha beta(+) T -ALL, less V beta-J beta 1 and more V beta-J beta 2 rearrangements were det ected in mature TCR alpha beta(+) T cell malignancies. The occurrence of th e different types of TCRB rearrangement patterns has implications for PCR-b ased clonality assessment and for PCR-based detection of minimal residual d isease via TCRB gene analysis. For instance, focussing on the beta 2 region of T-ALL will allow detection of rearrangements in 70%, 75%, and 90% of CD 3(-), TCR gamma delta(+), and TCR alpha beta(+) cases, respectively. Theref ore the here-described results will facilitate the design of the most appro priate primer sets for PCR analysis of TCRB gene rearrangements at the DNA level.