Primary antibodies used for immunocytochemistry were purchased from commercial suppliers: anti‐c‐FOS (4): sc‐52 (dilution: 1:200, Santa Cruz, #F0215) and anti‐c‐FOS (dilution: 1:1,000, Abcam, #ab208942). For Western blotting, the following primary antibodies were purchased from commercial suppliers: anti‐PSD95 (dilution 1:4,000, Synaptic Systems, #124011), anti‐Homer1 (dilution 1:10,000, Synaptic Systems, #160022), anti‐mGluR5 (dilution 1:1,000, Millipore, #2757164), and anti‐Beta III Tubulin (dilution 1:250,000, Covance, #PRB‐435P).
Anti mglur5
Manufactured by Merck Group
Sourced in United States
Anti-mGluR5 is a selective and potent antagonist of the metabotropic glutamate receptor 5 (mGluR5). It is used as a research tool to study the role of mGluR5 in various biological processes.
Automatically generated - may contain errors
3 protocols using anti mglur5
1
Immunocytochemistry and Western Blotting Protocols
2
Co-immunoprecipitation of mGluR5, ERα, and Homer
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Plasmid constructs were transfected or co-transfected into HEK293 using Lipofectamine 2000 reagent. 48 h post-transfection, cells were lysed in CoIP buffer (50 mm Tris, pH 7.4, 120 mm NaCl, 1 mM EDTA, 1% Triton X-100 containing protease and phosphase inhibitor mixture), and insoluble debris was removed by centrifugation (10 min, 10,000 g). The cleared lysate was incubated with an anti-Flag antibody (Sigma-Aldrich, F1804, 7 μg) or same IgG isotype control, while rotating, overnight at 4°C and then for 3–4 h with Dynabeads G beads. Bound proteins were eluted with sample buffer and loaded on gradient SDS-PAGE gels and processed for western blot as described above. Input lysate and CoIP complexes were blotted with anti-mGluR5 (Millipore, 1:50000 for input and 1:200000 for IP), anti-ERα antibodies (Millipore, 1:50000 for Input and 1:200000 for IP) and anti-Homer (Santa Cruz E18, 1:10000 for both IP and Input). GAPDH (Millipore, 1:300000) was used a loading control for input samples.
3
Antibody-based Detection of nAChR and GluR Subunits
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For the detection of nAChR subunits we used affinity-purified, subunit-specific, polyclonal antibodies (Abs), produced in rabbit against peptides derived from the intracytoplasmic loop (CYT) of nAChR subunits [27] . For the GluR subunits we produced Abs directed against the COOH peptide (EGYNVYGIESVKI) of the GluA2/3 subunit and of the Nterminal peptide (RTSDSRDHTRVDWKR) of the the GluA1 subunit. The specificity of the affinity-purified Abs was tested by western blotting studies using β2 WT and β2 KO mouse cortex extracts cDNAs (supplementary Fig. 1) or using cells transfected and non-transfected with GluA1, GluA2 and GluA3 cDNAs (supplementary Fig. 2).
The anti-GluN2B (clone N59/20), anti-PSD95 (clone K28/43) and anti-SAP102 (clone N19/2) Abs were from Antibodies incorporated (Davis, CA, USA); anti-GluN1 (clone 54.1) was from BD Pharmingen (Franklin Lakes, NJ, USA); anti-GluN2A (clone A3-2D10) was from Life technologies (Waltham, MA, USA); anti-mGluR5 was from Millipore (Billerica, MA, USA); anti-mGluR2 (clone mG2Na-s) was from Abcam (Cambridge, UK) and anti-actin (clone AC-40) was from Sigma-Aldrich (St. Louis, MO, USA).
The anti-GluN2B (clone N59/20), anti-PSD95 (clone K28/43) and anti-SAP102 (clone N19/2) Abs were from Antibodies incorporated (Davis, CA, USA); anti-GluN1 (clone 54.1) was from BD Pharmingen (Franklin Lakes, NJ, USA); anti-GluN2A (clone A3-2D10) was from Life technologies (Waltham, MA, USA); anti-mGluR5 was from Millipore (Billerica, MA, USA); anti-mGluR2 (clone mG2Na-s) was from Abcam (Cambridge, UK) and anti-actin (clone AC-40) was from Sigma-Aldrich (St. Louis, MO, USA).
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