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  • Biotechnical Methods Section BTS
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Biotechnical Methods Section (BTS)

Detection of hTERT protein by flow cytometry

Leukemia volume 14, pages 2176–2181 (2000)Cite this article

Abstract

Telomerase is a telomere-specific DNA polymerase consisting of protein and RNA components, which is activated in germline cells and the majority of cancers and serves to counter the consequences of telomere shortening. The protein component, hTERT, is believed to be the catalytic subunit of human telomerase and its expression at the mRNA level correlates well with telomerase activity in vitro. Current techniques for assaying telomerase activity detect only the mean activity in a sample and are unable to isolate specific cell sub-populations. This report describes the development and validation of a cellular, immunofluorescence-based flow cytometry assay that allows detection of intranuclear hTERT while maintaining identifiable cell population characteristics. The assay was shown to be both sensitive to changes in telomerase expression and was semi-quantitative. In both cell line differentiation experiments and in primary cells, a good correlation existed between hTERT expression measured by flow cytometry and telomerase activity detected by the telomeric repeat amplification protocol (TRAP). The method developed offers a quick, simple and reproducible cellular-based assay for hTERT expression. This assay will provide a useful, new tool for future investigations, facilitating the analysis of hTERT expression in mixed cell populations.

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Acknowledgements

ASM Ali is supported by the University of Manchester and the JHG Holt Scholarship and Special Exhibition in Medicine. R Chopra is supported by the Christie Hospital Leukaemia Endowment fund and Chugai Pharmaceuticals. J Robertson is supported by the Leukaemia Research Fund. NG Testa is supported by the Cancer Research Campaign.

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Authors and Affiliations

  1. Department of Experimental Haematology, Paterson Institute for Cancer Research, Manchester, UK

    ASM Ali, R Chopra, J Robertson & NG Testa

  2. Department of Medical Oncology, Paterson Institute for Cancer Research, Manchester, UK

    R Chopra

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  1. ASM Ali
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  2. R Chopra
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Ali, A., Chopra, R., Robertson, J. et al. Detection of hTERT protein by flow cytometry. Leukemia 14, 2176–2181 (2000). https://doi.org/10.1038/sj.leu.2401950

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