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Detection of hTERT protein by flow cytometry

Lookup NU author(s): Dr Ased Ali

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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.


Publication metadata

Author(s): Ali AS, Chopra R, Robertson J, Testa NG

Publication type: Article

Publication status: Published

Journal: Leukemia

Year: 2000

Volume: 14

Issue: 12

Pages: 2176-2181

Print publication date: 01/12/2000

ISSN (print): 0887-6924

ISSN (electronic): 1476-5551

Publisher: Nature Publishing Group

URL: http://dx.doi.org/10.1038/sj.leu.2401950

DOI: 10.1038/sj.leu.2401950


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