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Double In Situ Detection of Sonic Hedgehog mRNA and pMAPK Protein in Examining the Cell Proliferation Signaling Pathway in Mouse Embryo

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Book cover Signal Transduction Immunohistochemistry

Part of the book series: Methods in Molecular Biology ((MIMB,volume 717))

Abstract

Double in situ detection of RNA molecules and proteins in tissue sections is not trivial. A successful experiment heavily depends on the preparation of the tissue as well as the quality of the probes and antibodies. Detection of two or more molecular markers also requires reagents and experimental conditions that will preserve authenticity (accuracy) of the single staining patterns. Here, we describe in detail the protocols used to detect sonic hedgehog (Shh) mRNA by in situ hybridization and immunofluorescence staining for phosphorylated mitogen-activated protein kinase (pMAPK) in the same mouse embryonic tissue sections. In addition to protocols for manual immuno-staining, we provide data from automated machine-based staining protocols and highly recommend it to achieve strong signal and reproducible results.

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References

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Acknowledgments

We thank the members of the Molecular Cytology Core Facility for discussions and support and Dr. A.L. Joyner for Shh cDNA.

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Authors and Affiliations

  1. Developmental Biology Program, Molecular Cytology Core Facility, Memorial Sloan-Kettering Cancer Center, New York, NY, USA

    Sho Fujisawa, Mesruh Turkekul, Afsar Barlas, Ning Fan & Katia Manova

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  1. Sho Fujisawa
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  2. Mesruh Turkekul
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  3. Afsar Barlas
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  4. Ning Fan
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  5. Katia Manova
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Corresponding author

Correspondence to Katia Manova .

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Editors and Affiliations

  1. R & D Systems, Inc., McKinley Place NE. 614, Minneapolis, 55413, Minnesota, USA

    Alexander E. Kalyuzhny

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© 2011 Springer Science+Business Media, LLC

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Fujisawa, S., Turkekul, M., Barlas, A., Fan, N., Manova, K. (2011). Double In Situ Detection of Sonic Hedgehog mRNA and pMAPK Protein in Examining the Cell Proliferation Signaling Pathway in Mouse Embryo. In: Kalyuzhny, A. (eds) Signal Transduction Immunohistochemistry. Methods in Molecular Biology, vol 717. Humana Press. https://doi.org/10.1007/978-1-61779-024-9_15

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  • DOI: https://doi.org/10.1007/978-1-61779-024-9_15

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  • Publisher Name: Humana Press

  • Print ISBN: 978-1-61779-023-2

  • Online ISBN: 978-1-61779-024-9

  • eBook Packages: Springer Protocols

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