Rabbit polyclonal antibody to Mu Opioid Receptor (phospho Ser375).
Specificity
This antibody binds specifically to the phosphorylated form of human Mu Opioid Receptor at residue Ser375. It detects agonist-induced phosphorylation in Western blot analysis and supports subcellular localization by immunocytochemistry.
Applications
WB, ICC
Dilutions
WB: 1,000, ICC: 1:200
Reactivity
Human, Mouse, Rat
Immunogen
Synthetic phosphopeptide derived from human Mu Opioid Receptor around the phosphorylation site of Ser375. Immunogen range is 10-12 amino acids.
Host
Rabbit
Clonality
Polyclonal
Isotype
IgG
Conjugate
Unconjugated
Purification
Antigen affinity purification.
Concentration
Lot Specific
Product Form
Liquid
Formulation
Supplied in Dulbecco's PBS, pH 7.4, with 150 mM NaCl and 0.005% Sodium Azide.
Storage
Shipped at 4°C. Upon delivery aliquot and store at -20°C. Avoid freeze/thaw cycles.
Analysis of agonist-induced phosphorylation of Opioid Receptor Mu 1 (Mu Opioid Receptor) at Serine375. HEK293 cells stably expressing Mu Opioid Receptor were untreated or treated with 10 µM DAMGO or 0.1 µM PMA for 30 min. Lysates were immunoblotted with anti-Mu Opioid Receptor (phospho Ser375) antibody (A334520) at 1:1000 (upper panel). The blot was stripped and reprobed with phosphorylation-independent anti-Mu Opioid Receptor antibody at 1:1000 to confirm equal loading (lower panel).
Dose-dependent phosphorylation analysis of Opioid Receptor Mu 1 (Mu Opioid Receptor) using multiple phosphosite-specific antibodies. HEK293 cells stably expressing Mu Opioid Receptor were untreated or exposed to 1 nM to 1 µM DAMGO for 30 min. Lysates were immunoblotted with anti-Mu Opioid Receptor (phospho Thr370) (A334521), anti-Mu Opioid Receptor (phospho Ser375) (A334520), anti-Mu Opioid Receptor (phospho Thr376) (A334522), or anti-Mu Opioid Receptor (phospho Thr379) antibodies at 1:1000 (upper panels). The blot was stripped and reprobed with phosphorylation-independent anti-Mu Opioid Receptor antibody at 1:1000 to confirm equal loading (lower panel). Serine375 was identified as the primary phosphorylation site.
Agonist-selective phosphorylation analysis of Opioid Receptor Mu 1 (Mu Opioid Receptor) using multiple phosphosite-specific antibodies. HEK293 cells stably expressing Mu Opioid Receptor were untreated or exposed to 10 µM morphine or 10 µM DAMGO for 30 min. Lysates were immunoblotted with anti-Mu Opioid Receptor (phospho Thr370) (A334521), anti-Mu Opioid Receptor (phospho Ser375) (A334520), anti-Mu Opioid Receptor (phospho Thr376) (A334522), or anti-Mu Opioid Receptor (phospho Thr379) antibodies at 1:1000 (upper panels). The blot was stripped and reprobed with phosphorylation-independent anti-Mu Opioid Receptor antibody at 1:1000 to confirm equal loading (lower panel). DAMGO induced multisite phosphorylation, while morphine primarily induced Serine375 phosphorylation.
Immunocytochemical detection of Serine375 phosphorylation of Opioid Receptor Mu 1 (Mu Opioid Receptor). HEK293 cells stably expressing Mu Opioid Receptor were untreated or treated with 10 µM DAMGO for 30 min and stained with anti-Mu Opioid Receptor (phospho Ser375) antibody (A334520) at 1:200. Phosphorylated Mu Opioid Receptor receptors were undetectable in untreated cells (0 min) and observed at the plasma membrane and in perinuclear vesicular clusters after 30 min of DAMGO treatment.
In vivo detection of agonist-induced Serine375 phosphorylation of Opioid Receptor Mu 1 (Mu Opioid Receptor) in mouse brain. Mice were untreated or treated with 30 µg/kg etonitazene (subcutaneous). After 30 min, brain homogenates were immunoblotted with anti-Mu Opioid Receptor (phospho Ser375) antibody (A334520) at 1:1000 (upper panel). The blot was stripped and reprobed with phosphorylation-independent anti-Mu Opioid Receptor antibody at 1:1000 to confirm equal loading (lower panel). Phosphorylated Mu Opioid Receptor was detected only in wild-type (Mu Opioid Receptor-WT) mice after agonist treatment, absent in Mu Opioid Receptor-deficient (Mu Opioid Receptor-KO) mice.
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