Journal article
Characterization of Antibodies against Receptor Activity-Modifying Protein 1 (RAMP1): A Cautionary Tale
International journal of molecular sciences, Vol.23(24), p.16035
12/16/2022
DOI: 10.3390/ijms232416035
PMCID: PMC9787598
PMID: 36555690
Abstract
Calcitonin gene-related peptide (CGRP) is a key component of migraine pathophysiology, yielding effective migraine therapeutics. CGRP receptors contain a core accessory protein subunit: receptor activity-modifying protein 1 (RAMP1). Understanding of RAMP1 expression is incomplete, partly due to the challenges in identifying specific and validated antibody tools. We profiled antibodies for immunodetection of RAMP1 using Western blotting, immunocytochemistry and immunohistochemistry, including using RAMP1 knockout mouse tissue. Most antibodies could detect RAMP1 in Western blotting and immunocytochemistry using transfected cells. Two antibodies (844, ab256575) could detect a RAMP1-like band in Western blots of rodent brain but not RAMP1 knockout mice. However, cross-reactivity with other proteins was evident for all antibodies. This cross-reactivity prevented clear conclusions about RAMP1 anatomical localization, as each antibody detected a distinct pattern of immunoreactivity in rodent brain. We cannot confidently attribute immunoreactivity produced by RAMP1 antibodies (including 844) to the presence of RAMP1 protein in immunohistochemical applications in brain tissue. RAMP1 expression in brain and other tissues therefore needs to be revisited using RAMP1 antibodies that have been comprehensively validated using multiple strategies to establish multiple lines of convincing evidence. As RAMP1 is important for other GPCR/ligand pairings, our results have broader significance beyond the CGRP field.
Details
- Title: Subtitle
- Characterization of Antibodies against Receptor Activity-Modifying Protein 1 (RAMP1): A Cautionary Tale
- Creators
- Erica R Hendrikse - University of AucklandTayla A Rees - University of AucklandZoe Tasma - University of AucklandMichael L Garelja - University of OtagoAndrew Siow - University of AucklandPaul W R Harris - University of AucklandJohn B Pawlak - University of North Carolina at Chapel HillKathleen M Caron - University of North Carolina at Chapel HillElizabeth S Blakeney - University of North Carolina at Chapel HillAndrew F Russo - Center for the Prevention and Treatment of Visual Loss, Veterans Administration Health Center, Iowa City, IA 52246, USALevi P Sowers - Center for the Prevention and Treatment of Visual Loss, Veterans Administration Health Center, Iowa City, IA 52246, USAThomas A Lutz - University of ZurichChristelle Le Foll - University of ZurichChristopher S Walker - University of AucklandDebbie L Hay - University of Auckland
- Resource Type
- Journal article
- Publication Details
- International journal of molecular sciences, Vol.23(24), p.16035
- DOI
- 10.3390/ijms232416035
- PMID
- 36555690
- PMCID
- PMC9787598
- NLM abbreviation
- Int J Mol Sci
- ISSN
- 1422-0067
- eISSN
- 1422-0067
- Grant note
- RF1NS113839 / NIH HHS
- Language
- English
- Date published
- 12/16/2022
- Academic Unit
- Neurology; Molecular Physiology and Biophysics; Stead Family Department of Pediatrics; Iowa Neuroscience Institute; Craniofacial Anomalies Research Center; Neurology (Pediatrics)
- Record Identifier
- 9984339314002771
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