Human/Mouse/Rat GPR64 Antibody

Detection of Rat and Mouse GPR64 by Western Blot. Western blot shows lysates of rat epididymis tissue and mouse epididymis tissue. PVDF membrane was probed with 2 µg/mL of Sheep Anti-Human/Mouse/Rat GPR64 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF7977) followed by HRP-conjugated Anti-Sheep IgG Secondary Antibody (Catalog # HAF016). A specific band was detected for GPR64 at approximately 150 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

GPR64 in Human Epididymus. GPR64 was detected in immersion fixed paraffin-embedded sections of human epididymus using Sheep Anti-Human/Mouse/Rat GPR64 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF7977) at 3 µg/mL overnight at 4 °C. Before incubation with the primary antibody, tissue was subjected to heat-induced epitope retrieval using Antigen Retrieval Reagent-Basic (Catalog # CTS013). Tissue was stained using the Anti-Sheep HRP-DAB Cell & Tissue Staining Kit (brown; Catalog # CTS019) and counterstained with hematoxylin (blue). Specific staining was localized to plasma membranes of epithelial cells. View our protocol for Chromogenic IHC Staining of Paraffin-embedded Tissue Sections.

Detection of GPR64 in HEK293 Human Cell Line Transfected with Human GPR64 and eGFP by Flow Cytometry. HEK293 human embryonic kidney cell line transfected with (A) Human GPR64 or (B) Human GPR75 and eGFP was stained with Sheep Anti-Human GPR64 Affinity-Purified Polyclonal Antibody (Catalog # AF7977) followed by Allophycocyanin-conjugated Anti-Sheep IgG Secondary Antibody (Catalog # F0127). Quadrant markers were set based on control antibody staining (Catalog # 5-001-A). View our protocol for Staining Membrane-associated Proteins.

Detection of GPR64 by Western Blot The expression of ADGRG2, CFTR, Gs, Gq, beta -arrestin-1, beta -arrestin-2 in efferent ductules, brain and liver tissue of WT and Adgrg2-/Y mice.(A) Western blot analysis of ADGRG2, CFTR, Gs, Gq, beta -arrestin-1, beta -arrestin-2 expression in efferent ductules, brain and liver tissue of WT and Adgrg2-/Y mice. A representative western blot from at least three independent experiments was shown. (CFTR antibody:20738–1-AP, Proteintech). (B) Bar graph representation and statistical analyses of (A). All blots were normalized to GAPDH. n.s., no significant difference; Adgrg2-/Y mice compared with WT mice in the same tissue. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/29393851), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of GPR64 by Western Blot The complex formation between ADGRG2, beta -arrestin-1 and CFTR in HEK293 cells.(A) HEK293 cells were transfected with equal amount plasmids encoding ADGRG2, CFTR, beta -arrestin-1 or beta -arrestin-2 plasmids. The Flag- ADGRG2 were pulled down by M2-Flag beads and the associated CFTR, beta -arrestins were detected by western blot. Representative images from at least three independent experiments are shown (CFTR antibody:20738–1-AP, Proteintech). (B) Bar graph representation and statistical analyses of (A).***p<0.001, IP protein of ADGRG2 overexpressed cells were compared with control plasmids transfected cells respectively. n.s., no significant difference. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/29393851), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of GPR64 by Western Blot The ADGRG2 protein knockout strategy, PCR strategy and western blot results of Adgrg2-/Y and Gnaq+/- mice.(A) Schematic representation of the ADGRG2 knockout strategy for the Adgrg2-/Y mice. 2 bp nucleotides were removed and 10 bp nucleotides were inserted in the first exon of the ADGRG2 gene in the ADGRG2 mutant mice by the CRISP-CAS9 approach. In the ADGRG2-deficient mice, the translation of ADGRG2 was terminated at the 7th amio acid after the signal peptide. (B) Schematic representation for the primers used in the genotyping of the ADGRG2 mutant mice or their wild-type littermates. (C) Schematic description of the PCR strategy and expected results for genotyping. The genotyping of mice was determined by PCR and visualized by bromide staining of agarose Gels. (D) Western blot analysis of ADGRG2 expression in efferent duct tissue of WT and Adgrg2-/Y mice. All blots were normalized to GAPDH. (E) Bar graph representation and statistical analyses of (D). At least three independent experiments were carried out. (F) Western blot analysis of Gq expression in efferent duct tissue of WT and Gnaq+/- mice. All blots were normalized to GAPDH. (G) Bar graph representation and statistical analyses of (F). At least three independent experiments were carried out. (E,G) *p<0.05, **p<0.01, ***p<0.001, Adgrg2-/Y mice or Gnaq+/- mice were compared with WT mice. n.s., no significant difference. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/29393851), licensed under a CC-BY license. Not internally tested by R&D Systems.