Overview and assessment of the histochemical methods and reagents for the detection of 尾-galactosidase activity in transgenic animals

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• 作者：Stefan Trifonov ; Yuji Yamashita ; Masahiko Kase…
• 关键词：尾 ; galactosidase ; lacZ ; Nitroblue tetrazolium ; Salmon ; gal ; X ; gal
• 刊名：Anatomical Science International
• 出版年：2016
• 出版时间：January 2016
• 年：2016
• 卷：91
• 期：1
• 页码：56-67
• 全文大小：7,730 KB
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• 作者单位：Stefan Trifonov (1)
  Yuji Yamashita (1)
  Masahiko Kase (1)
  Masato Maruyama (1)
  Tetsuo Sugimoto (1)
  
  1. Department of Anatomy and Brain Science, Kansai Medical University, 2-5-1 Shin-machi, Hirakata, Osaka, 573-1010, Japan
  
• 刊物主题：Anatomy; Human Physiology; Animal Physiology; Cell Biology; Animal Anatomy / Morphology / Histology;
• 出版者：Springer Japan
• ISSN：1447-073X

文摘

Bacterial 尾-galactosidase is one of the most widely used reporter genes in experiments involving transgenic and knockout animals. In this review we discuss the current histochemical methods and available reagents to detect 尾-galactosidase activity. Different substrates are available, but the most commonly used is X-gal in combination with potassium ferri- and ferro-cyanide. The reaction produces a characteristic blue precipitate in the cells expressing 尾-galactosidase, and despite its efficiency in staining whole embryos, its detection on thin tissue sections is difficult. Salmon-gal is another substrate, which in combination with ferric and ferrous ions gives a reddish-pink precipitate. Its sensitivity for staining tissue sections is similar to that of X-gal. Combining X-gal or Salmon-gal with tetrazolium salts provides a faster and more sensitive reaction than traditional 尾-galactosidase histochemistry. Here, we compare the traditional 尾-galactosidase assay and the combination of X-gal or Salmon-gal with three tetrazolium salts: nitroblue tetrazolium, tetranitroblue tetrazolium and iodonitrotetrazolium. Based on an assessment of the sensitivity and specificity of the different combinations of substrates, we are proposing an optimized and enhanced method for 尾-galactosidase detection in histological sections of the transgenic mouse brain. Optimal staining was obtained with X-gal in combination with nitroblue tetrazolium, which provides a faster and more specific staining than the traditional X-gal combination with potassium ferri- and ferro-cyanide. We recommend the X-gal/nitroblue tetrazolium staining mixture as the first choice for the detection of 尾-galactosidase activity on histological sections. When faster results are needed, Salmon-gal/nitroblue tetrazolium should be considered as an alternative, while maintaining acceptable levels of noise. Keywords 尾-galactosidase lacZ Nitroblue tetrazolium Salmon-gal X-gal