Detection of T cell receptor circles (TRECs) as biomarkers for de novo T cell synthesis using a quantitative polymerase chain reaction-enzyme linked immunosorbent assay (PCR-ELISA)

Lena Al-Harthi, Giulia Marchetti, Carolyn M. Steffens, Jean François Poulin, Rafick Pierre Sékaly, Alan Landay

Research output: Contribution to journalArticlepeer-review

72 Scopus citations

Abstract

Currently, phenotypic markers that distinguish between recent thymic emigrants/de novo T cells and the rest of the peripheral T cell pool are lacking. This distinction is critical in studies aimed at evaluating immune reconstitution following intensive chemotherapy, in immunodeficiency-related therapies, or in the elucidation of the kinetics of thymic function. During V(D)J T cell receptor rearrangement, DNA extrachromosomal excision products are generated. These products, known as T cell receptor excision circles (TRECs), are not replicated during mitosis and are thus diluted with each round of cell division. Therefore, TRECs can be used as an indicator of recent thymic emigrants. Thus far, quantitative competitive-polymerase chain reaction (QC-PCR) and real time PCR were used to measure TREC levels. However, QC-PCR relies on radioactivity, is cumbersome when processing many samples at once and the cost of real time PCR does not make it a viable option for many laboratories. We describe here the development of a quantitative PCR-ELISA method for the measurement of coding joint TRECs generated from VαJα recombination. Our assay is ultra sensitive, relies on biotin labeling rather than radioactivity, is based on a 96-well format making multiple process sampling relatively easy, and is cost effective. Using this PCR-ELISA method, we evaluated thymic output among 22 normal subjects, ranging in age from 22-53 years, and among HIV-infected individuals following highly active antiretroviral therapy (HAART). We demonstrate that an inverse relationship exists between TREC levels and aging in normal individuals and that, among some HIV patients, HAART treatment leads to enhanced thymic output. Our assay has direct relevance in projects examining normal and abnormal thymic function and in immune reconstitution studies. Copyright (C) 2000 Elsevier Science B.V.

Original languageEnglish (US)
Pages (from-to)187-197
Number of pages11
JournalJournal of Immunological Methods
Volume237
Issue number1-2
DOIs
StatePublished - Apr 3 2000
Externally publishedYes

Keywords

  • De novo T cells
  • PCR-ELISA
  • Recent thymic emigrants
  • T cell receptor excision circle (TREC)
  • Thymopoiesis

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology

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