Open AccessDetection and Characterization of αβ-T-Cell Clonality by Denaturing Gradient Gel Electrophoresis (DGGE)
Author(s) -
Per thor Straten,
Annette Malene Barfoed,
Tina Seremet,
Ingvil Sæterdal,
Jesper Zeuthen,
Per Guldberg
Publication year - 1998
Publication title -
biotechniques/biotechniques
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.617
H-Index - 131
eISSN - 1940-9818
pISSN - 0736-6205
DOI - 10.2144/98252st05
Subject(s) - temperature gradient gel electrophoresis , biology , polyclonal antibodies , gel electrophoresis , t cell receptor , microbiology and biotechnology , t cell , cell , gene , antigen , genetics , immune system , 16s ribosomal rna
Accumulation of T cells carrying identical T-cell receptors (TCR) is associated with a number of immunological and nonimmunological diseases. Therefore, it is of interest to be able to analyze complex T-cell populations for the presence of clonally expanded subpopulations. Here, we describe a simple method combining reverse transcription (RT)-PCR and denaturing gradient gel electrophoresis (DGGE) for rapid detection and characterization of T-cell clonality. The detection of clonally expanded 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. For polyclonal populations with a high degree of junctional diversity, the different DNA molecules will resolve at different positions in the gel and together will be revealed as a smear. For each of the TCR β-variable gene (BV)1–24 families, cloned transcripts were amplified and shown to resolve as distinct bands in the denaturing gradient gel, whereas the analysis 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 monitoring T-cell responses in diagnostic and therapeutic settings.