Immunohistochemical Detection of Sphingosine-1-Phosphate and Sphingosine Kinase-1 in Human Tissue Samples and Cell Lines

  • Gary M. Reynolds
  • Barbara Visentin
  • Roger Sabbadini
Part of the Methods in Molecular Biology book series (MIMB, volume 1697)


Sphingosine-1-phosphate (S1P) and the enzyme primarily responsible for its production, sphingosine kinase-1 (SphK-1), are dysregulated in multiple human diseases including cancer, multiple sclerosis (MS), diabetes, neurological diseases, fibrosis, and certain pathologies associated with impaired angiogenesis such as age-related macular degeneration (AMD). Antibody-based techniques to identify and localize S1P and SphK-1 within cells and tissue specimens represent a powerful tool, not only to understand biological role of these molecules but also to validate these unique in-class targets in multiple state diseases. Consequently, the potential applications of these molecules for therapy and diagnostic purposes are currently under investigation. Here, we describe a new improved technique, Agitated Low Temperature Epitope Retrieval (ALTER) for staining procedures, to identify expression of S1P and SphK-1 in human frozen tissue samples. The challenges encountered in the process of localization in tissue samples of lipid molecules such as S1P are discussed.


Bioactive lipids S1P SPHK1 Anti-S1P antibody LT002 ALTER technique Immunohistochemistry Liver cancer 


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Copyright information

© Springer Science+Business Media New York 2017

Authors and Affiliations

  • Gary M. Reynolds
    • 1
  • Barbara Visentin
    • 2
  • Roger Sabbadini
    • 3
  1. 1.Centre for Liver Research and NIHR Biomedical Research UnitUniversity of Birmingham and Queen Elizabeth Hospital BirminghamBirminghamUK
  2. 2.La Jolla Biologics IncSan DiegoUSA
  3. 3.Stanford University School of MedicineStanfordUSA

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