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Characterization of a human telomerase reverse transcriptase sequence containing two antigenic epitopes with high affinity for human leucocyte antigen
Author(s) -
Wang Jinwen,
Yu Liang,
Li Jianfeng,
Deng Riqiang,
Wang Xunzhang
Publication year - 2007
Publication title -
biotechnology and applied biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.468
H-Index - 70
eISSN - 1470-8744
pISSN - 0885-4513
DOI - 10.1042/ba20060245
Subject(s) - telomerase reverse transcriptase , epitope , antigen , biology , telomerase , reverse transcriptase , microbiology and biotechnology , polyclonal antibodies , antibody , virology , polymerase chain reaction , immunology , gene , biochemistry
Almost all human malignant tumours exhibit strong telomerase activity, but normal adult tissues, with a few exceptions, do not. hTERT (human telomerase reverse transcriptase) is an essential component of telomerase, and hence it can serve as a parallel sign in the diagnosis and prognosis of cancers. In the present study, we selected a sequence of hTERT containing two antigenic epitopes that have high affinity for HLA‐A2 (human leucocyte antigen‐A2) as a TAA (tumour‐associated antigen) based on a peptide–motif scoring system. The sequence was obtained by reverse‐transcriptase PCR and cloned into the Escherichia coli expression vector pGEX‐4T‐1. The expression product appeared in the form of inclusion bodies. Denatured inclusion‐body extract was subjected to SDS/PAGE, and the gel band corresponding to the putative 38 kDa fusion protein (GST–hTERT major tumour‐associated antigen) was excised, ground with PBS, mixed with Freund's adjuvant and used to inoculate mice, generating anti‐TERT polyclonal antibodies. Western blotting using the leukaemia cell line THP‐1 demonstrated that the antibodies were able to detect hTERT expression, implying the potential applicability of the antigenic peptides derived from hTERT as a universal marker in the diagnosis and prognosis of tumours.