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

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 beta 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 beta chain complementary DNA is amplified by a nested polymerase chain reaction with V beta family-specific oligonucleotide primers and a fluorochrome-labeled C beta 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.

journal_name

Am J Clin Pathol

authors

Lue C,Mitani Y,Crew MD,George JF,Fink LM,Schichman SA

doi

10.1093/ajcp/111.5.683

subject

Has Abstract

pub_date

1999-05-01 00:00:00

pages

683-90

issue

5

eissn

0002-9173

issn

1943-7722

journal_volume

111

pub_type

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