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Immunohistochemistry to Study YAP in Human Tissue Samples

  • Franziska Haderk
  • Victor Olivas
  • Trever G. Bivona
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1893)

Abstract

Immunohistochemistry (IHC) analysis of YAP in human tissue samples represents an important means to analyze overall expression levels and subcellular localization of YAP in specimen of interest. As transcriptional coactivator, alterations of YAP levels in the cellular nucleus allow important predictions for YAP activity and transcriptional state of target genes. In the following report, IHC procedures optimized for the detection of YAP in tissue slides of FFPE material are provided. Of note, de-paraffinization and heat-induced antigen retrieval are strictly necessary for successful YAP IHC staining. Further, immunostaining using a labelled polymer-HRP system combined with diaminobenzidine (DAB), as signal-amplifying chromogen, allows strong staining results with minimal unspecific background signal.

Key words

YAP Immunohistochemistry IHC YAP nuclear activity FFPE samples Hippo signaling Digital pathology 

Notes

Acknowledgments

The authors acknowledge funding from NIH/NCI R01CA211052 and NIH/NCI R01CA204302 (to T.G.B.) as well as from the German Cancer Aid (Mildred Scheel postdoctoral fellowship, to F.H.).

References

  1. 1.
    Huang J, Wu S, Barrera J et al (2005) The Hippo signaling pathway coordinately regulates cell proliferation and apoptosis by inactivating Yorkie, the Drosophila homolog of YAP. Cell 122:421–434CrossRefGoogle Scholar
  2. 2.
    Moroishi T, Hansen CG, Guan K-L (2015) The emerging roles of YAP and TAZ in cancer. Nat Rev Cancer 15:73–79CrossRefGoogle Scholar
  3. 3.
    Zhao B, Wei X, Li W et al (2007) Inactivation of YAP oncoprotein by the Hippo pathway is involved in cell contact inhibition and tissue growth control. Genes Dev 21:2747–2761CrossRefGoogle Scholar
  4. 4.
    Zhao B, Ye X, Yu J et al (2008) TEAD mediates YAP-dependent gene induction and growth control. Genes Dev 22:1962–1971CrossRefGoogle Scholar
  5. 5.
    Lin L, Sabnis AJ, Chan E et al (2015) The Hippo effector YAP promotes resistance to RAF- and MEK-targeted cancer therapies. Nat Genet 47:250–256CrossRefGoogle Scholar
  6. 6.
    McGowan M, Kleinberg L, Halvorsen AR et al (2017) NSCLC depend upon YAP expression and nuclear localization after acquiring resistance to EGFR inhibitors. Genes Cancer 8:497–504PubMedPubMedCentralGoogle Scholar
  7. 7.
    Liu G, Yu F-X, Kim YC et al (2015) Kaposi sarcoma-associated herpesvirus promotes tumorigenesis by modulating the Hippo pathway. Oncogene 34:3536–3546CrossRefGoogle Scholar
  8. 8.
    Yang S, Zhang L, Purohit V et al (2015) Active YAP promotes pancreatic cancer cell motility, invasion and tumorigenesis in a mitotic phosphorylation-dependent manner through LPAR3. Oncotarget 6:36019–36031PubMedPubMedCentralGoogle Scholar
  9. 9.
    Nakane PK, Pierce GB (1966) Enzyme-labeled antibodies: preparation and application for the localization of antigens. J Histochem Cytochem 14:929–931CrossRefGoogle Scholar
  10. 10.
    Huang SN, Minassian H, More JD (1976) Application of immunofluorescent staining on paraffin sections improved by trypsin digestion. Lab Invest 35:383–390PubMedGoogle Scholar
  11. 11.
    Hsu SM, Raine L, Fanger H (1981) Use of avidin-biotin-peroxidase complex (ABC) in immunoperoxidase techniques: a comparison between ABC and unlabeled antibody (PAP) procedures. J Histochem Cytochem 29:577–580CrossRefGoogle Scholar
  12. 12.
    Singer SJ (1959) Preparation of an electron-dense antibody conjugate. Nature 183:1523–1524CrossRefGoogle Scholar
  13. 13.
    Haines DM, West KH (2005) Immunohistochemistry: forging the links between immunology and pathology. Vet Immunol Immunopathol 108:151–156CrossRefGoogle Scholar
  14. 14.
    de Matos LL, Trufelli DC, de Matos MGL et al (2010) Immunohistochemistry as an important tool in biomarkers detection and clinical practice. Biomark Insights 5:9–20CrossRefGoogle Scholar
  15. 15.
    Taylor CR, Rudbeck L (2013) Immunohistochemical Staining Methods. Dako Denmark A/SGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Franziska Haderk
    • 1
    • 2
    • 3
  • Victor Olivas
    • 1
    • 2
    • 3
  • Trever G. Bivona
    • 1
    • 2
    • 3
  1. 1.Department of MedicineUniversity of California, San FranciscoSan FranciscoUSA
  2. 2.Department of Cellular and Molecular PharmacologyUniversity of California, San FranciscoSan FranciscoUSA
  3. 3.Helen Diller Family Comprehensive Cancer CenterUniversity of California, San FranciscoSan FranciscoUSA

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