Heat-assisted stretching of paraffin sections on hot plate weakens immunoreactivity of orotate phosphoribosyltransferase

Shingo Kamoshida, Nao Sakamoto, Hiroshi Matsuoka, Yoichi Sakurai, Kazuki Sakamoto, Yoshikazu Sugimoto, Masakazu Fukushima, Yutaka Tsutsumi

研究成果: Article査読

6 被引用数 (Scopus)

抄録

Orotate phosphoribosyltransferase (OPRT) is the key enzyme for the phosphorylation of 5-fluorouracil (5-FU), the rate-limiting step for acquiring its anti-tumor effect Since high enzyme activities and rnRNA levels of OPRT are said to be associated with 5-FU chemosensitivity of cancer cells, the immunohistochemical demonstration of OPRT can be expected to contribute to the selection of patients who suffer from 5-FU-sensitive cancer. During a study for establishing the appropriate immunostaining condition using rabbit antiserum in formalin-fixed, paraffin-embedded sections of human cancer, we unexpectedly uncovered the fact that heat-assisted stretching of paraffin sections on a hot plate just after sectioning was critical for preserving OPRT immunoreactivity; namely, stretching sections briefly at 70°C or higher significantly reduced the immunoreactivity. Overnight drying of the sections in an oven at 37°C or 60°C did not influence the immunoreactivity, but pretreatments, including 0.2% trypsin, 0.002% proteinase K, and pressure cooking in 10 mM citrate buffer, pH 6.0 and pH 7.0, and 1 mM ethylenediaminetetraacetic acid solution, pH 8.0, seriously deteriorated the OPRT epitopes. The immunoreactivity was relatively resistant to overfixation, though weakened after fixation in formalin for four weeks. Xenografts with high OPRT enzyme activities showed distinct positive cytoplasmic staining, but those with low OPRT activities were equivocal. OPRT expression in routinely processed cancer tissues also provided valuable data.

本文言語English
ページ(範囲)69-74
ページ数6
ジャーナルActa Histochemica et Cytochemica
38
1
DOI
出版ステータスPublished - 2005
外部発表はい

ASJC Scopus subject areas

  • Pathology and Forensic Medicine
  • Biochemistry
  • Physiology
  • Histology
  • Cell Biology

フィンガープリント 「Heat-assisted stretching of paraffin sections on hot plate weakens immunoreactivity of orotate phosphoribosyltransferase」の研究トピックを掘り下げます。これらがまとまってユニークなフィンガープリントを構成します。

引用スタイル