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Detection of recombinant and endogenous mouse melatonin receptors by monoclonal antibodies targeting the C‐terminal domain
Author(s) -
Cecon Erika,
Ivanova Anna,
Luka Marine,
Gbahou Florence,
Friederich Anne,
Guillaume JeanLuc,
Keller Patrick,
Knoch Klaus,
Ahmad Raise,
Delagrange Philippe,
Solimena Michele,
Jockers Ralf
Publication year - 2019
Publication title -
journal of pineal research
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.881
H-Index - 131
eISSN - 1600-079X
pISSN - 0742-3098
DOI - 10.1111/jpi.12540
Subject(s) - melatonin , melatonin receptor , receptor , monoclonal antibody , biology , immunofluorescence , endocrinology , microbiology and biotechnology , antibody , medicine , immunology , biochemistry
Melatonin receptors play important roles in the regulation of circadian and seasonal rhythms, sleep, retinal functions, the immune system, depression, and type 2 diabetes development. Melatonin receptors are approved drug targets for insomnia, non‐24‐hour sleep‐wake disorders, and major depressive disorders. In mammals, two melatonin receptors (MTRs) exist, MT 1 and MT 2 , belonging to the G protein‐coupled receptor (GPCR) superfamily. Similar to most other GPCRs, reliable antibodies recognizing melatonin receptors proved to be difficult to obtain. Here, we describe the development of the first monoclonal antibodies (mABs) for mouse MT 1 and MT 2 . Purified antibodies were extensively characterized for specific reactivity with mouse, rat, and human MT 1 and MT 2 by Western blot, immunoprecipitation, immunofluorescence, and proximity ligation assay. Several mABs were specific for either mouse MT 1 or MT 2 . None of the mABs cross‐reacted with rat MTRs, and some were able to react with human MTRs. The specificity of the selected mABs was validated by immunofluorescence microscopy in three established locations (retina, suprachiasmatic nuclei, pituitary gland) for MTR expression in mice using MTR‐KO mice as control. MT 2 expression was not detected in mouse insulinoma MIN6 cells or pancreatic beta‐cells. Collectively, we report the first monoclonal antibodies recognizing recombinant and native mouse melatonin receptors that will be valuable tools for future studies.