Next Article in Journal
Role of Groucho and Groucho1-like in Regulating Metamorphosis and Ovary Development in Nilaparvata lugens (Stål)
Next Article in Special Issue
GPR18-Mediated Relaxation of Human Isolated Pulmonary Arteries
Previous Article in Journal
Meta-Analysis of APP Expression Modulated by SARS-CoV-2 Infection via the ACE2 Receptor
Previous Article in Special Issue
Membrane Melatonin Receptors Activated Cell Signaling in Physiology and Disease
 
 
Article

Suitability of GRK Antibodies for Individual Detection and Quantification of GRK Isoforms in Western Blots

Institut für Molekulare Zellbiologie, CMB—Center for Molecular Biomedicine, Universitätsklinikum Jena, Friedrich-Schiller-Universität Jena, Hans-Knöll-Straße 2, D-07745 Jena, Germany
*
Author to whom correspondence should be addressed.
Academic Editor: Alessandro Cannavo
Int. J. Mol. Sci. 2022, 23(3), 1195; https://doi.org/10.3390/ijms23031195
Received: 26 December 2021 / Revised: 17 January 2022 / Accepted: 18 January 2022 / Published: 21 January 2022
G protein-coupled receptors (GPCRs) are regulated by GPCR kinases (GRKs) which phosphorylate intracellular domains of the active receptor. This results in the recruitment of arrestins, leading to desensitization and internalization of the GPCR. Aside from acting on GPCRs, GRKs regulate a variety of membrane, cytosolic, and nuclear proteins not only via phosphorylation but also by acting as scaffolding partners. GRKs’ versatility is also reflected by their diverse roles in pathological conditions such as cancer, malaria, Parkinson’s-, cardiovascular-, and metabolic disease. Reliable tools to study GRKs are the key to specify their role in complex cellular signaling networks. Thus, we examined the specificity of eight commercially available antibodies targeting the four ubiquitously expressed GRKs (GRK2, GRK3, GRK5, and GRK6) in Western blot analysis. We identified one antibody that did not recognize its antigen, as well as antibodies that showed unspecific signals or cross-reactivity. Hence, we strongly recommend testing any antibody with exogenously expressed proteins to clearly confirm identity of the obtained Western blot results. Utilizing the most-suitable antibodies, we established the Western blot-based, cost-effective simple tag-guided analysis of relative protein abundance (STARPA). This method allows comparison of protein levels obtained by immunoblotting with different antibodies. Furthermore, we applied STARPA to determine GRK protein levels in nine commonly used cell lines, revealing differential isoform expression. View Full-Text
Keywords: GRK2; GRK3; GRK5; GRK6; antibody specificity; Western blot; protein quantification GRK2; GRK3; GRK5; GRK6; antibody specificity; Western blot; protein quantification
Show Figures

Figure 1

MDPI and ACS Style

Reichel, M.; Weitzel, V.; Klement, L.; Hoffmann, C.; Drube, J. Suitability of GRK Antibodies for Individual Detection and Quantification of GRK Isoforms in Western Blots. Int. J. Mol. Sci. 2022, 23, 1195. https://doi.org/10.3390/ijms23031195

AMA Style

Reichel M, Weitzel V, Klement L, Hoffmann C, Drube J. Suitability of GRK Antibodies for Individual Detection and Quantification of GRK Isoforms in Western Blots. International Journal of Molecular Sciences. 2022; 23(3):1195. https://doi.org/10.3390/ijms23031195

Chicago/Turabian Style

Reichel, Mona, Verena Weitzel, Laura Klement, Carsten Hoffmann, and Julia Drube. 2022. "Suitability of GRK Antibodies for Individual Detection and Quantification of GRK Isoforms in Western Blots" International Journal of Molecular Sciences 23, no. 3: 1195. https://doi.org/10.3390/ijms23031195

Find Other Styles
Note that from the first issue of 2016, MDPI journals use article numbers instead of page numbers. See further details here.

Article Access Map by Country/Region

1
Back to TopTop