Protocol to characterize G(i/o) and G(s) protein-coupled receptors in transiently transfected cells using ELISA and cAMP measurements

Activation of G(s) or G(i/o) protein-coupled receptors (GPCRs) leads to changes of intracellular cyclic adenosine monophosphate (cAMP) levels. This protocol describes steps for cloning HA- and FLAG-tagged GPCRs, transient transfection of CHO-K1 or HEK293-T cells, and determination of basal and ligan...

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Autores principales: Liebing, Aenne-Dorothea, Krumbholz, Petra, Stäubert, Claudia
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Elsevier 2023
Materias:
Protocol
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9958081/
https://www.ncbi.nlm.nih.gov/pubmed/36853674
http://dx.doi.org/10.1016/j.xpro.2023.102120
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author Liebing, Aenne-Dorothea
Krumbholz, Petra
Stäubert, Claudia
author_facet Liebing, Aenne-Dorothea
Krumbholz, Petra
Stäubert, Claudia
author_sort Liebing, Aenne-Dorothea
collection PubMed
description Activation of G(s) or G(i/o) protein-coupled receptors (GPCRs) leads to changes of intracellular cyclic adenosine monophosphate (cAMP) levels. This protocol describes steps for cloning HA- and FLAG-tagged GPCRs, transient transfection of CHO-K1 or HEK293-T cells, and determination of basal and ligand-induced changes in intracellular cAMP levels. We detail enzyme-linked immunosorbent assays to determine relative GPCR plasma membrane and total expression levels. For complete details on the use and execution of this protocol, please refer to Schulze et al. (2022).(1)
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spelling pubmed-99580812023-02-26 Protocol to characterize G(i/o) and G(s) protein-coupled receptors in transiently transfected cells using ELISA and cAMP measurements Liebing, Aenne-Dorothea Krumbholz, Petra Stäubert, Claudia STAR Protoc Protocol Activation of G(s) or G(i/o) protein-coupled receptors (GPCRs) leads to changes of intracellular cyclic adenosine monophosphate (cAMP) levels. This protocol describes steps for cloning HA- and FLAG-tagged GPCRs, transient transfection of CHO-K1 or HEK293-T cells, and determination of basal and ligand-induced changes in intracellular cAMP levels. We detail enzyme-linked immunosorbent assays to determine relative GPCR plasma membrane and total expression levels. For complete details on the use and execution of this protocol, please refer to Schulze et al. (2022).(1) Elsevier 2023-02-14 /pmc/articles/PMC9958081/ /pubmed/36853674 http://dx.doi.org/10.1016/j.xpro.2023.102120 Text en © 2023 The Author(s) https://creativecommons.org/licenses/by-nc-nd/4.0/This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
spellingShingle Protocol
Liebing, Aenne-Dorothea
Krumbholz, Petra
Stäubert, Claudia
Protocol to characterize G(i/o) and G(s) protein-coupled receptors in transiently transfected cells using ELISA and cAMP measurements
title Protocol to characterize G(i/o) and G(s) protein-coupled receptors in transiently transfected cells using ELISA and cAMP measurements
title_full Protocol to characterize G(i/o) and G(s) protein-coupled receptors in transiently transfected cells using ELISA and cAMP measurements
title_fullStr Protocol to characterize G(i/o) and G(s) protein-coupled receptors in transiently transfected cells using ELISA and cAMP measurements
title_full_unstemmed Protocol to characterize G(i/o) and G(s) protein-coupled receptors in transiently transfected cells using ELISA and cAMP measurements
title_short Protocol to characterize G(i/o) and G(s) protein-coupled receptors in transiently transfected cells using ELISA and cAMP measurements
title_sort protocol to characterize g(i/o) and g(s) protein-coupled receptors in transiently transfected cells using elisa and camp measurements
topic Protocol
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9958081/
https://www.ncbi.nlm.nih.gov/pubmed/36853674
http://dx.doi.org/10.1016/j.xpro.2023.102120
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