Nuclear telomerase is less accessible to antibody probing than known nuclear antigens: retrieval with new immunostaining buffer.
Telomerase is an important tumor marker but few antibodies to the enzyme have been described or used without difficulty in histochemical detection. Here we report specific detection of the enzyme in cell and tissue preparations using a new monoclonal antibody (mAb 476) and a new antigen-retrieval buffer (Enhancing buffer). When used to detect telomerase under normal immunostaining conditions in HL-60 cells or tissue sections of hepatocellular carcinoma or metastatic choriocarcinoma, unexpectedly, the antibody stained the cytoplasm rather than the nucleus. Nuclear staining, however, was revealed using the Enhancing buffer. Since other nuclear antigens in the HL-60 cell could be stained both ordinarily and in the Enhancing buffer, nuclear telomerase appears to be shrouded by the nuclear matrix or blocked by accessory proteins. The cytoplasmic activity seen in normal buffer but absent largely from the Enhancing buffer may be an artifact or the nascent, "naked" enzyme. With a known cytoplasmic antigen (proteinase-3) chosen arbitrarily for comparison, the antigenicity was found enhanced, instead, by the Enhancing buffer. The mode of action of the Enhancing buffer differs from that of microwave irradiation or the signal amplification (CSA) used by some investigators. The latter was found to enhance the cytoplasmic reactivity rather than the nuclear reactivity of mAb 476.
Leung
Danny Tze-Ming
DT
Clinical Immunology Unit, Prince of Wales Hospital, The Chinese University of Hong Kong, Shatin, Hong Kong. pllim@cuhk.edu.hk
Ma
Chun-Hung
CH
Niu
Haitao
H
Liew
Choong-Tsek
CT
Tang
Janet Tsui-Ying
JT
Lim
Pak-Leong
PL
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000918:Antibody Specificity; D014408:Biomarkers, Tumor / Q000276:immunology*; D002021:Buffers; D006528:Carcinoma, Hepatocellular / Q000201:enzymology / Q000473:pathology; D002467:Cell Nucleus / Q000201:enzymology* / Q000276:immunology; D002822:Choriocarcinoma / Q000201:enzymology / Q000556:secondary; D003593:Cytoplasm / Q000201:enzymology / Q000276:immunology; D000939:Epitopes; D018922:HL-60 Cells; D006801:Humans; D007150:Immunohistochemistry; D007414:Intestinal Neoplasms / Q000201:enzymology / Q000556:secondary; D008113:Liver Neoplasms / Q000201:enzymology / Q000473:pathology; D051379:Mice; D008807:Mice, Inbred BALB C; D012333:RNA, Messenger; D019098:Telomerase / Q000235:genetics / Q000276:immunology*
D000911:Antibodies, Monoclonal; D014408:Biomarkers, Tumor; D002021:Buffers; D000939:Epitopes; D012333:RNA, Messenger; D019098:Telomerase