An automated method for the analysis of T-cell receptor repertoires: Rapid RT-PCR fragment length analysis of the T-cell receptor chain complementarity-determining region 3

Academic Article

Abstract

  • The examination of T-cell receptor (TCR) repertoires has an important role in the study of lymphoproliferative disorders and autoimmune diseases. Analysis of the complementarity-determining region 3 (CDR3) of the TCR β chain is used to assess the clonality of T-cell populations. We developed a rapid fluorescence-based method for CDR3 length analysis of expressed TCR gene families. TCR β chain complementary DNA is amplified by a nested polymerase chain reaction with Vβ family-specific oligonucleotide primers and a fluorochrome-labeled Cβ primer. The polymerase chain reaction products were analyzed on a compact automated DNA sequencing system (OpenGene system, Visible Genetics, Toronto, Ontario). To demonstrate the usefulness of our technique, we examined the CDR3 length distribution of peripheral blood T cells from a healthy subject, intestinal T cells from a patient with ulcerative colitis, and the T-cell leukemia cell line Jurkat. The analysis revealed polyclonal, oligoclonal, and monoclonal CDR3 distributions, respectively, for the 3 T-cell populations. Our new method shows virtually identical CDR3 length patterns compared with the traditional radioisotope- based method. The new technique offers the convenience of rapid throughput, nonradioactive labeling, and quality data analysis.
  • Authors

    Digital Object Identifier (doi)

    Author List

  • Lue C; Mitani Y; Crew MD; George JF; Fink LM; Schichman SA
  • Start Page

  • 683
  • End Page

  • 690
  • Volume

  • 111
  • Issue

  • 5