Analysis of T cell receptor-γ gene rearrangements by denaturing gradient gel electrophoresis of GC-clamped polymerase chain reaction products: Correlation with tumor-specific sequences

We describe a modified denaturing gradient gel electrophoresis (DGGE) procedure with a 40-nucleotide GC clamp in the polymerase chain reaction to improve resolution in amplifying T cell receptor-γ (TCR-γ) rearrangements. DNA from 46 cases of lymphoblastic leukemia/lymphoma, 5 T cell lines, 2 B cell lines, 7 normal lymphocytes, and 3 cases of Hodgkin's disease was amplified by polymerase chain reaction. In addition, 20 cases of paraffin- embedded T cell lymphomas and 5 cases of reactive hyperplasia were also studied. Clonal TCR-γ rearrangements were identified on DGGE by the presence of a predominant band. Results obtained from 5 T cell lines and 12 lymphoblastic leukemia/lymphomas containing known TCR-γ gene rearrangements revealed 100% concordance in detecting clonal rearrangements between DGGE and traditional Southern blot analysis. Of the remaining 34 lymphoblastic leukemia/lymphoma cases studied by DGGE alone, 30 were positive. DGGE analysis of 10 lymphoblastic leukemia/lymphoma cases with known group IVγ to Jγ1 or Jγ2 rearrangement sequences confirmed that the electrophoretic migration was dependent on the tumor-specific rearranged TCR-γ sequence. In addition, 17 of 20 cases of paraffin-embedded T cell lymphomas were positive by DGGE, 6 of which had the clonal population also identified in fresh tissue DNA. DGGE analysis of GC-clamped polymerase chain reaction products can provide a way to more accurately detect TCR-γ clonality of lymphoid tumors and can be applied to archival tissues.