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Detection of the human GPR50 orphan seven transmembrane protein by polyclonal antibodies mapping different epitopes


Hamouda, H O; Chen, P; Levoye, A; Sözer-Topçular, N; Daulat, A M; Guillaume, J L; Ravid, R; Savaskan, E; Ferry, G; Boutin, J A; Delagrange, P; Jockers, R; Maurice, P (2007). Detection of the human GPR50 orphan seven transmembrane protein by polyclonal antibodies mapping different epitopes. Journal of Pineal Research, 43(1):10-15.

Abstract

GPR50 is an orphan seven transmembrane protein related to the melatonin receptor subfamily comprising MT(1) and MT(2) receptors. In the absence of any known ligand for GPR50, other tools are critical for the characterization of this protein. Here, we describe the generation, purification and characterization of the first rabbit polyclonal antibodies generated against peptides corresponding to the N-terminus, C-terminus and two additional regions within the intracellular tail of GPR50. Immune sera were purified on peptide-antigen affinity columns. Antibodies specifically recognized a GPR50-YFP fusion protein on the plasma membrane of HEK 293 cells in immunofluorescence experiments. In Western blot experiments, the monomeric and dimeric forms of GPR50 were detected as proteins of 66 and 130 kDa, respectively. In addition, these new antibodies were sufficiently sensitive to detect GPR50 in brain slices of the rat pituitary and human hippocampus. In conclusion, we successfully produced antibodies against the orphan GPR50 protein that will become valuable tools for functional studies of this protein.

GPR50 is an orphan seven transmembrane protein related to the melatonin receptor subfamily comprising MT(1) and MT(2) receptors. In the absence of any known ligand for GPR50, other tools are critical for the characterization of this protein. Here, we describe the generation, purification and characterization of the first rabbit polyclonal antibodies generated against peptides corresponding to the N-terminus, C-terminus and two additional regions within the intracellular tail of GPR50. Immune sera were purified on peptide-antigen affinity columns. Antibodies specifically recognized a GPR50-YFP fusion protein on the plasma membrane of HEK 293 cells in immunofluorescence experiments. In Western blot experiments, the monomeric and dimeric forms of GPR50 were detected as proteins of 66 and 130 kDa, respectively. In addition, these new antibodies were sufficiently sensitive to detect GPR50 in brain slices of the rat pituitary and human hippocampus. In conclusion, we successfully produced antibodies against the orphan GPR50 protein that will become valuable tools for functional studies of this protein.

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Additional indexing

Item Type:Journal Article, refereed, original work
Communities & Collections:04 Faculty of Medicine > Psychiatric University Hospital Zurich > Division of Psychiatric Research and Clinic for Psychogeriatric Medicine
Dewey Decimal Classification:610 Medicine & health
Language:English
Date:2007
Deposited On:06 Sep 2011 08:30
Last Modified:05 Apr 2016 14:59
Publisher:Wiley-Blackwell
ISSN:0742-3098
Publisher DOI:10.1111/j.1600-079X.2007.00437.x
PubMed ID:17614830

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