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:10.1371/journal.pone.0020606

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

Research output: Contribution to journal › Article › peer-review

27 Citations (Scopus)

Abstract

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.

Original language	English
Article number	e20606
Journal	PLoS ONE
Volume	6
Issue number	8
DOIs	https://doi.org/10.1371/journal.pone.0020606
Publication status	Published - 1 Jan 2011
Externally published	Yes

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SDG 3 Good Health and Well-being

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10.1371/journal.pone.0020606Licence: CC BY

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Kasprowicz, V. O., Mitchell, J. E., Chetty, S., Govender, P., Huang, K. H. G., Fletcher, H. A., Webster, D. P., Brown, S., Kasmar, A., Millington, K., Day, C. L., Mkhwanazi, N., McClurg, C., Chonco, F., Lalvani, A., Walker, B. D., Ndung'u, T., & Klenerman, P. (2011). A molecular assay for sensitive detection of pathogen-specific T-cells. PLoS ONE, 6(8), Article e20606. https://doi.org/10.1371/journal.pone.0020606

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title = "A molecular assay for sensitive detection of pathogen-specific T-cells",

abstract = "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.",

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

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Kasprowicz, VO, Mitchell, JE, Chetty, S, Govender, P, Huang, KHG, Fletcher, HA, Webster, DP, Brown, S, Kasmar, A, Millington, K, Day, CL, Mkhwanazi, N, McClurg, C, Chonco, F, Lalvani, A, Walker, BD, Ndung'u, T & Klenerman, P 2011, 'A molecular assay for sensitive detection of pathogen-specific T-cells', PLoS ONE, vol. 6, no. 8, e20606. https://doi.org/10.1371/journal.pone.0020606

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AU - Kasprowicz, Victoria O.

AU - Mitchell, Jessica E.

AU - Chetty, Shivan

AU - Govender, Pamla

AU - Huang, Kuan Hsiang Gary

AU - Fletcher, Helen A.

AU - Webster, Daniel P.

AU - Brown, Sebastian

AU - Kasmar, Anne

AU - Millington, Kerry

AU - Day, Cheryl L.

AU - Mkhwanazi, Nompumelelo

AU - McClurg, Cheryl

AU - Chonco, Fundisiwe

AU - Lalvani, Ajit

AU - Walker, Bruce D.

AU - Ndung'u, Thumbi

AU - Klenerman, Paul

PY - 2011/1/1

Y1 - 2011/1/1

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AB - 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.

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DO - 10.1371/journal.pone.0020606

M3 - Article

VL - 6

JO - PLoS ONE

JF - PLoS ONE

IS - 8

M1 - e20606

ER -

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

Abstract

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