PLoS ONE (Jan 2011)

A molecular assay for sensitive detection of pathogen-specific T-cells.

  • Victoria O Kasprowicz,
  • Jessica E Mitchell,
  • Shivan Chetty,
  • Pamla Govender,
  • Kuan-Hsiang Gary Huang,
  • Helen A Fletcher,
  • Daniel P Webster,
  • Sebastian Brown,
  • Anne Kasmar,
  • Kerry Millington,
  • Cheryl L Day,
  • Nompumelelo Mkhwanazi,
  • Cheryl McClurg,
  • Fundisiwe Chonco,
  • Ajit Lalvani,
  • Bruce D Walker,
  • Thumbi Ndung'u,
  • Paul Klenerman

DOI
https://doi.org/10.1371/journal.pone.0020606
Journal volume & issue
Vol. 6, no. 8
p. e20606

Abstract

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Here we describe the development and validation of a highly sensitive assay of antigen-specific IFN-γ production using real time quantitative PCR (qPCR) for two reporters--monokine-induced by IFN-γ (MIG) and the IFN-γ inducible protein-10 (IP10). We developed and validated the assay and applied it to the detection of CMV, HIV and Mycobacterium tuberculosis (MTB) specific responses, in a cohort of HIV co-infected patients. We compared the sensitivity of this assay to that of the ex vivo RD1 (ESAT-6 and CFP-10)-specific IFN-γ Elispot assay. We observed a clear quantitative correlation between the two assays (P<0.001). Our assay proved to be a sensitive assay for the detection of MTB-specific T cells, could be performed on whole blood samples of fingerprick (50 uL) volumes, and was not affected by HIV-mediated immunosuppression. This assay platform is potentially of utility in diagnosis of infection in this and other clinical settings.