In Situ Hybridization for RNA: Nonradioactive Probe: Oligo-DNA Probe: Digoxigenin (1)

  • Yuka Iwasaki
  • Ihachi Kato
Part of the Springer Lab Manuals book series (SLM)


In this article, protocols are described for preparation of digoxigenin-labeled oligo-DNA probe and their use in in situ hybridization. As has been shown previously (Koji et al. 1988, 1989; Koji and Brenner 1993; Koji and Nakane 1990, 1996), nonradioactive synthetic oligo-DNA probes can be very useful tools. The 3’ -end of oligo-DNA can be labeled enzymaticallywith several possible haptens, such as digoxigenin and biotin. Digoxigenin, a plant steroidal aglycone, is especially suitable for immunocytochemical detection because its epitope is not present in animal cells. Also, it has recently been confirmed that under appropriate conditions, there is no substantial difference in the sensitivity of detection between [35S] cRNA (Denjin et al. 1990) or [35S] cDNA (Unger et al. 1991) probes and nonradioactive probes.


Dextran Sulfate Keratinocyte Growth Factor Chromogen Solution Cell BioI Nonradioactive Probe 
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  1. Adams J.C. (1981) Heavy metal intensification of DAB-based HRP reaction product. J Histochem Cytochem 29: 775PubMedCrossRefGoogle Scholar
  2. Denjin M, De Weger R.A., Berends M.J.H, Compier-Spies P.I., Jansz H, Van Unnik J.A.M., Lips C.J.M. (1990) Detection of calcitonin-encoding mRNA by radioactive and non-radioactive in situ hybridization: improved colorimetric detection and cellular localization of mRNA in thyroid sections. J Histochem Cytochem 38: 351–358CrossRefGoogle Scholar
  3. Iwasaki Y, Hara Y, Koji T, Shibata Y, Nakane P.K., Kato I (1998) Differential expression of IFN-γ,IL-4, IL-10, and IL-1ß mRNAs in decalcified tissue sections of mouse lipopolysaccharide-induced periodontitis mandibles assessed by in situ hybridization.Histochem and Cell Biol1090:339–347CrossRefGoogle Scholar
  4. Koji T, Brenner R.M. (1993) Localization of estrogen receptor messenger ribonucleic acid in rhesus monkey uterus by nonradioactive in situ hybridization with digoxigenin-labeled oligodeoxynucleotides. Endocrinology 132: 382–392PubMedCrossRefGoogle Scholar
  5. Koji T, Nakane P.K. (1990) Localization in situ of specific mRNA using thymine-thymine dimmerized DNA probes. Sensitive and reliable nonradioactive in situ hybridization. Acta Pathol Jpn 40: 793–807PubMedGoogle Scholar
  6. Koji T, Nakane P.K. (1996) Recent advances in molecular histochemical techniques: in situ hybridization and southwestern histochemistry. J Electron Microsc 45: 119–127CrossRefGoogle Scholar
  7. Koji T, Chedid M, Rubin J.S., Slayden O.D., Csaky K.G., Aaronson S.A., Brenner R.M. (1994) Progesterone-dependent expression of keratinocyte growth factor mRNA in stromal cells of the primate endometrium: keratinocyte growth factor as a progestomedin. J Cell Biol 125: 393–401PubMedCrossRefGoogle Scholar
  8. Koji T, Moriuchi T, Nakane P.K. (1988) Improved tissue preparation for in situ localization of specific mRNA using non-radioactive DNA probes: effects of protease digestion and probe size on signal detection in frozen and paraffin sections of rat pituitary glands. Acta Histochem Cytochem 21: 187–200CrossRefGoogle Scholar
  9. Koji T, Sugawara I, Kimura M, Nakane P.K. (1989) In situ localization of c-myc mRNA in HL-60 cells using non-radioactive synthetic oligodeoxynucleotide probes. Acta Histochem Cytochem 22: 295–307CrossRefGoogle Scholar
  10. Unger E.R., Hammer M.L., Chenggis M.L. (1991) Comparison of 35S and biotin as labels for in situ hybridization: use of an HPV model system. J Histochem Cytochem 39: 145–150PubMedCrossRefGoogle Scholar
  11. Yoshii A, Koji T, Ohsawa N, Nakane P.K. (1995) In situ localization of ribosomal RNAs is a reliable reference for hybridizable RNA in tissue sections.J Histochem Cytochem 43: 321–327PubMedCrossRefGoogle Scholar

Copyright information

© Springer Japan 2000

Authors and Affiliations

  • Yuka Iwasaki
    • 1
  • Ihachi Kato
  1. 1.Department of PeriodontologyNagasaki University School of DentistryNagasakiJapan

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