No answer to the lack of specificity: mouse monoclonal antibody targeting the angiotensin II type 1 receptor AT1 fails to recognize its target

No answer to the lack of specificity: mouse monoclonal antibody targeting the angiotensin II type... Numerous antibodies targeting G protein-coupled receptors (GPCRs) have been described as non-specific among the polyclonal antibodies against angiotensin II type 1 receptor (AT1). We have tested the newly developed AT1 receptor mouse monoclonal antibody for its specificity. Human embryonic kidney (HEK293) cells, which do not endogenously express AT1 receptor, were transfected in order to overexpress a fluorescently labeled enhanced green fluorescent protein (EGFP)-tagged human AT1 receptor. Western blot and immunofluorescence assays were performed to test the specificity of the Santa Cruz monoclonal antibody sc-57036. These results were compared to the ones obtained with the polyclonal sc-1173 anti-AT1 receptor antibodies that have already been described as non-specific. While the positive controls using GFP antibodies detected the EGFP-tagged AT1 receptor, both polyclonal and monoclonal anti-AT1 receptor antibodies failed to specifically recognize the corresponding band by Western blot, as similar bands were revealed in either transfected or non-transfected cells. It also failed to detect AT1 receptor in immunofluorescence experiments. The lack of target recognition of the monoclonal AT1 receptor antibody in our experimental conditions suggests that this antibody could give misleading results such as misidentification of the protein. To our knowledge, no specific antibodies targeting AT1 receptors have been developed so far and the field is thus in need of new technical developments. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Naunyn-Schmiedeberg's Archives of Pharmacology Springer Journals

No answer to the lack of specificity: mouse monoclonal antibody targeting the angiotensin II type 1 receptor AT1 fails to recognize its target

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Publisher
Springer Berlin Heidelberg
Copyright
Copyright © 2018 by Springer-Verlag GmbH Germany, part of Springer Nature
Subject
Biomedicine; Pharmacology/Toxicology; Neurosciences
ISSN
0028-1298
eISSN
1432-1912
D.O.I.
10.1007/s00210-018-1522-4
Publisher site
See Article on Publisher Site

Abstract

Numerous antibodies targeting G protein-coupled receptors (GPCRs) have been described as non-specific among the polyclonal antibodies against angiotensin II type 1 receptor (AT1). We have tested the newly developed AT1 receptor mouse monoclonal antibody for its specificity. Human embryonic kidney (HEK293) cells, which do not endogenously express AT1 receptor, were transfected in order to overexpress a fluorescently labeled enhanced green fluorescent protein (EGFP)-tagged human AT1 receptor. Western blot and immunofluorescence assays were performed to test the specificity of the Santa Cruz monoclonal antibody sc-57036. These results were compared to the ones obtained with the polyclonal sc-1173 anti-AT1 receptor antibodies that have already been described as non-specific. While the positive controls using GFP antibodies detected the EGFP-tagged AT1 receptor, both polyclonal and monoclonal anti-AT1 receptor antibodies failed to specifically recognize the corresponding band by Western blot, as similar bands were revealed in either transfected or non-transfected cells. It also failed to detect AT1 receptor in immunofluorescence experiments. The lack of target recognition of the monoclonal AT1 receptor antibody in our experimental conditions suggests that this antibody could give misleading results such as misidentification of the protein. To our knowledge, no specific antibodies targeting AT1 receptors have been developed so far and the field is thus in need of new technical developments.

Journal

Naunyn-Schmiedeberg's Archives of PharmacologySpringer Journals

Published: Jun 4, 2018

References

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