DETECTION AND CHARACTERIZATION OF ALPHA-BETA-T-CELL CLONALITY BY DENATURING GRADIENT GEL-ELECTROPHORESIS (DGGE)

Accumulation of T cells carrying identical T-cell,receptors (TCR) is a ssociated with a number of immunological and nonimmunological diseases . Therefore, it is of interest to be able to analyze complex T-cell po pulations for the presence of clonally expanded subpopulations. Here, we describe a simple method combining reverse transcription (RT)-PCR a nd denaturing gradient gel electrophoresis (DCGE) for rapid detection and characterization of T-ceIl clonality The detection of clonality ex panded T cells by DGGE relies on the fact that clonal transcripts have no junctional diversity and therefore resolve at a fixed position in the gel,,which is determined by their melting properties. I;br polyclo nal populations with a high degree of junctional diversity the differe nt DNA molecules will resolve at different positions in the gel and to gether,will be revealed a smear. For each of the TCR beta-variable gen e (BV)1-24 families, cloned transcripts were amplified and shown to re solve as distinct bands in the denaturing gradient gel,,whereas the an alysis of polyclonal T-cell populations resulted in a smear in the gel . The present method might prove useful to test for clonotypic T-cells in a variety of pathological and physiological conditions and for, mo nitoring T-cell responses in diagnostic and therapeutic settings.