Abstract
AlphaScreen® SureFire® assay is a novel technology that combines luminescent oxygen channeling technology, nano-beads, and monocloncal antibodies to detect the level of a selected protein in a volume lower than 5 μl. This method is more sensitive compared with the traditional enzyme-linked immunosorbent assays (ELISA), and can detect an increasing number of new targets. Here, we described a method for AlphaScreen® SureFire® assay that targets ERK1/2 phosphorylation, a primary downstream signaling pathway that conveys activation of GPR55 by l-α-lysophosphatidylinositol (LPI) and certain cannabinoids.
This is a preview of subscription content, log in via an institution.
Buying options
Tax calculation will be finalised at checkout
Purchases are for personal use only
Learn about institutional subscriptionsSpringer Nature is developing a new tool to find and evaluate Protocols. Learn more
References
Anavi-Goffer S, Baillie G, Irving AJ et al (2012) Modulation of L-alpha-lysophosphatidylinositol/GPR55 mitogen-activated protein kinase (MAPK) signaling by cannabinoids. J Biol Chem 287:91–104
Kotsikorou E, Sharir H, Shore DM et al (2013) Identification of the GPR55 antagonist binding site using a novel set of high-potency GPR55 selective ligands. Biochemistry 52:9456–9469
Heynen-Genel S, Dahl R, Shi S, et al (2010) Screening for selective ligands for GPR55 agonists in Probe Reports from the NIH Molecular Libraries Program (Internet), National Center for Biotechnology Information, Bethesda, MD, NBK66152 [bookaccession]
Henstridge CM, Balenga NA, Ford LA et al (2009) The GPR55 ligand L-alpha-lysophosphatidylinositol promotes RhoA-dependent Ca2+ signaling and NFAT activation. FASEB J 23:183–193
Andradas C, Caffarel MM, Perez-Gomez E et al (2011) The orphan G protein-coupled receptor GPR55 promotes cancer cell proliferation via ERK. Oncogene 30:245–252
Ford LA, Roelofs AJ, Anavi-Goffer S et al (2010) A role for L-alpha-lysophosphatidylinositol and GPR55 in the modulation of migration, orientation and polarization of human breast cancer cells. Br J Pharmacol 160:762–771
Pineiro R, Maffucci T, Falasca MT (2011) The putative cannabinoid receptor GPR55 defines a novel autocrine loop in cancer cell proliferation. Oncogene 30:142–152
Heynen-Genel S, Dahl R, Shi S, et al (2010) Screening for selective ligands for GPR55 antagonists in Probe Reports from the NIH Molecular Libraries Program (Internet), National Center for Biotechnology Information, Bethesda, MD, NBK66153 [bookaccession]
Staton PC, Hatcher JP, Walker DJ et al (2008) The putative cannabinoid receptor GPR55 plays a role in mechanical hyperalgesia associated with inflammatory and neuropathic pain. Pain 139:225–236
Shih HH, Zhang S, Cao W et al (2009) CRP is a novel ligand for the oxidized LDL receptor LOX-1. Am J Physiol Heart Circ Physiol 296:H1643–H1650
Acknowledgments
This work was supported by the US National Institutes of Health grant numbers DA-03672 and DA-09789 (RAR). We thank the University of Aberdeen for a Knowledge Transfer Grant Award.
Conflicts of interest: RAR received funding from GW Pharmaceuticals.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2016 Springer Science+Business Media New York
About this protocol
Cite this protocol
Anavi-Goffer, S., Ross, R.A. (2016). A Functional Assay for GPR55: Envision Protocol. In: Maccarrone, M. (eds) Endocannabinoid Signaling. Methods in Molecular Biology, vol 1412. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3539-0_8
Download citation
DOI: https://doi.org/10.1007/978-1-4939-3539-0_8
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-3537-6
Online ISBN: 978-1-4939-3539-0
eBook Packages: Springer Protocols