Detection of clonal T-cell receptor gamma gene rearrangements using fluorescent-based PCR and automated high-resolution capillary electrophoresis

Background: Analysis of T-cell receptor gamma (TCR gamma) gene rearrangemen ts by PCR is a powerful tool for detecting clonal T-cell populations for th e diagnosis of lymphoid neoplasms. We report a method for TCR gamma PCR ana lysis using capillary electrophoresis (CE). Methods and Results: To define the threshold for identification of a predom inant monoclonal population within a polyclonal background, we developed a novel objective parameter of the peak height ratio (Rn) of the peak of inte rest and the average of the two immediate flanking peaks. After evaluation of monoclonal, reactive, and normal T-cell populations, an Rn of 3.0 or gre ater was determined to be consistent with a monoclonal population, whereas an Rn between 1.9 and 3.0 was considered an intermediate range. This CE met hod was compared with the standard denaturing gradient gel electrophoresis (DGGE) method using previously evaluated clinical specimens. Eleven of 12 c linical specimens (92%) with a definitive diagnosis of T-cell lymphoma. wer e monoclonal by CE, with 100% concordance with the DGGE method. Of nine spe cimens morphologically suspicious for T-cell lymphoma, five specimens were positive by CE analysis compared with four specimens by DGGE. In addition, 14 specimens for staging from patients with known T-cell lymphoma were stud ied using both the CE and DGGE methods, with a concordance of 86%. Conclusion: CE is a powerful and efficient method for analysis of clonality by TCR gamma PCR.