Characterization of Antibodies against Receptor Activity-Modifying Protein 1 (RAMP1): A Cautionary Tale
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Hendrikse, E.R, et al. Characterization of Antibodies Against Receptor Activity-modifying Protein 1 (ramp1): A Cautionary Tale. MDPI, 2022. https://doi.org/10.17615/x8gt-a573APA
Hendrikse, E., Rees, T., Tasma, Z., Garelja, M., Siow, A., Harris, P., Pawlak, J., Caron, K., Blakeney, E., Russo, A., Sowers, L., Lutz, T., Le Foll, C., Walker, C., & Hay, D. (2022). Characterization of Antibodies against Receptor Activity-Modifying Protein 1 (RAMP1): A Cautionary Tale. MDPI. https://doi.org/10.17615/x8gt-a573Chicago
Hendrikse, E.R., T.A Rees, Z Tasma, M.L Garelja, A Siow, P.W.R Harris, J.B Pawlak et al. 2022. Characterization of Antibodies Against Receptor Activity-Modifying Protein 1 (ramp1): A Cautionary Tale. MDPI. https://doi.org/10.17615/x8gt-a573- Creator
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Hendrikse, E.R.
- Other Affiliation: The University of Auckland
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Rees, T.A.
- Other Affiliation: The University of Auckland
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Tasma, Z.
- Other Affiliation: The University of Auckland
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Garelja, M.L.
- Other Affiliation: The University of Otago
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Siow, A.
- Other Affiliation: The University of Auckland
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Harris, P.W.R.
- Other Affiliation: The University of Auckland
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Pawlak, J.B.
- Affiliation: School of Medicine, Department of Cell Biology and Physiology
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Caron, K.M.
- Affiliation: School of Medicine, Department of Cell Biology and Physiology
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Blakeney, E.S.
- Affiliation: School of Medicine, Department of Cell Biology and Physiology
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Russo, A.F.
- Other Affiliation: University of Iowa
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Sowers, L.P.
- Other Affiliation: University of Iowa
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Lutz, T.A.
- Other Affiliation: University of Zurich
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Le Foll, C.
- Other Affiliation: University of Zurich
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Walker, C.S.
- Other Affiliation: The University of Auckland
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Hay, D.L.
- Other Affiliation: The University of Auckland
- 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.
- Date of publication
- 2022
- Keyword
- DOI
- Identifier
- Resource type
- Article
- Rights statement
- In Copyright
- License
- Attribution 4.0 International
- Journal title
- International Journal of Molecular Sciences
- Journal volume
- 23
- Journal issue
- 24
- Language
- English
- Version
- Publisher
- Funder
- National Institutes of Health, NIH: RF1NS113839; National Institute of Neurological Disorders and Stroke, NINDS; Health Research Council of New Zealand, HRC; Auckland Medical Research Foundation, AMRF; University of Auckland; Schweizerischer Nationalfonds zur Förderung der Wissenschaftlichen Forschung, SNF: SNF310030_ 207763; Maurice Wilkins Centre for Molecular Biodiscovery
- ISSN
- 1661-6596
- Publisher
- MDPI
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