Silent Synapses Antibody Sampler Kit
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Catalog# HAK21066
Silent Synapses Antibody Sampler Kit
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WB
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Human
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Mouse
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Rat
General Overview
Kit Components
| Product Includes | Specification | Application | Reactivity | Mw |
|---|---|---|---|---|
| Glutamate receptor 1[ET1612-10] | 20µl | WB,IHC-P,IP,IHC-Fr | Human,Mouse,Rat | Predicted band size: 102 kDa |
| Phospho-GluR1 (S845)[ET1701-28] | 20µl | WB,FC | Mouse,Rat | Predicted band size: 102 kDa |
| Phospho-GluR1 (AMPA subtype) (S831)[HA721865] | 20µl | WB | Human,Mouse | Predicted band size: 102 kDa |
| GluR2[ET1706-06] | 20µl | WB,IF-Cell,IHC-P | Human,Mouse,Rat | Predicted band size: 99 kDa |
| PSD95[ET1602-20] | 20µl | WB,IF-Cell,IF-Tissue,IHC-P,FC,IP,IHC-Fr | Human,Mouse,Rat | Predicted band size: 80 kDa |
| NMDAR1[ET1703-75] | 20µl | WB,IF-Cell,IF-Tissue,IHC-P,FC,IHC-Fr | Human,Mouse,Rat | Predicted band size: 105 kDa |
| Goat Anti-Rabbit IgG (H+L)[HA1001] | 100µl | WB,ELISA,IHC-P | Rabbit |
Product Description
The Silent Synapses Antibody Sampler Kit provides an economical means of detecting the activation of AMPA-type glutamate receptors (AMPAR) using phospho-specific and control antibodies. AMPARs expression can be compared to other synaptic components including NMDA-type glutamate receptor subunit GluN1 and the synaptic scaffolding protein PSD95. The kit includes enough antibody to perform two western blot experiments with each primary antibody.
Product Features
Storage Buffer
1*TBS (pH7.4), 0.05% BSA, 40% Glycerol. Preservative: 0.05% Sodium Azide.
Storage Instructions
Store at +4℃ after thawing. Aliquot store at -20℃. Avoid repeated freeze / thaw cycles.
Background
GluA2-containing AMPARs, AMPARs that lack GluA2 are permeable to calcium.Post-transcriptional modifications (alternative splicing, nuclear RNA editing) and post-translational modifications (glycosylation, phosphorylation) result in a very large number of permutations, fine-tuning the kinetic properties and surface expression of AMPARs representing key pathways to mediate synaptic plasticity. <BR>During development and mature states, some synapses exhibit “silent synapses” that lack functional AMPAR-mediated transmission.<BR>Synapses become “unsilenced” by post-translational modification of GluAs, particularly GluA1, which alters its kinetic properties and/or surface expression while other synaptic components, such as other glutamate receptors like NMDARs and postsynaptic scaffolding proteins like PSD95, remain unaltered.Conversely, reducing the AMPAR kinetic properties and surface expression can silence synapses.Key post-translational modifications implicated in regulating these processes include phosphorylation of GluA1 at Ser831 and Ser845.
Data Links
Background References
1. Diering GH, Heo S, Hussain NK, Liu B, Huganir RL. Extensive phosphorylation of AMPA receptors in neurons. Proc Natl Acad Sci U S A. 2016 Aug 16;113(33):E4920-7.
2. Meyers AM, Gnazzo F, Barrera ED, Nabatian T, Chan L, Beeler J. DIETARY REGULATION OF SILENT SYNAPSES IN THE DORSOLATERAL STRIATUM. bioRxiv [Preprint]. 2024 Mar 27:2024.03.24.586457.
Images
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Western blot analysis of Glutamate receptor 1 on different lysates with Rabbit anti-Glutamate receptor 1 antibody (ET1612-10) at 1/1,000 dilution.
Lane 1: Human brain tissue lysate (40 µg/Lane)
Lane 2: Mouse brain tissue lysate (40 µg/Lane)
Lane 3: Mouse cerebellum tissue lysate (40 µg/Lane)
Lane 4: Rat brain tissue lysate (40 µg/Lane)
Lane 5: Rat cerebellum tissue lysate (40 µg/Lane)
Predicted band size: 90 kDa
Observed band size: 90 kDa
Exposure time: 2 minutes
4-20% SDS-PAGE gel.
Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (ET1612-10) at 1/1,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature. -
Western blot analysis of Phospho-GluR1 (S845) on different lysates with Rabbit anti-Phospho-GluR1 (S845) antibody (ET1701-28) at 1/1,000 dilution.
Lane 1: Mouse brain tissue lysate (40 µg/Lane)
Lane 2: Rat brain tissue lysate (40 µg/Lane)
Lane 3: Mouse brain tissue lysate, the membrane treated with λpp for 1 hour (40 µg/Lane)
Lane 4: Rat brain tissue lysate, the membrane treated with λpp for 1 hour (40 µg/Lane)
Predicted band size: 102 kDa
Observed band size: 102 kDa
Exposure time: 3 minutes; ECL: K1801;
4-20% SDS-PAGE gel.
Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (ET1701-28) at 1/1,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature. -
Western blot analysis of PSD95 on different lysates with Rabbit anti-PSD95 antibody (ET1602-20) at 1/2,000 dilution.
Lane 1: Human brain tissue lysate (20 µg/Lane)
Lane 2: Mouse brain tissue lysate (20 µg/Lane)
Lane 3: Mouse hippocampus tissue lysate (20 µg/Lane)
Lane 4: Rat brain tissue lysate (20 µg/Lane)
Lane 5: Rat hippocampus tissue lysate (20 µg/Lane)
Predicted band size: 80 kDa
Observed band size: 75-100 kDa
Exposure time: 20 seconds; ECL: K1801;
4-20% SDS-PAGE gel.
Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (ET1602-20) at 1/2,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature. -
Western blot analysis of NMDAR1 on different lysates with Rabbit anti-NMDAR1 antibody (ET1703-75) at 1/5,000 dilution.
Lane 1: MCF7 cell lysate (15 µg/Lane)
Lane 2: Human brain tissue lysate (20 µg/Lane)
Lane 3: Mouse brain tissue lysate (20 µg/Lane)
Lane 4: Rat brain tissue lysate (20 µg/Lane)
Lane 5: Mouse heart tissue lysate (negative) (20 µg/Lane)
Lane 6: Rat liver tissue lysate (negative) (20 µg/Lane)
Predicted band size: 105 kDa
Observed band size: 120 kDa
Exposure time: 1 minute 2 seconds;
4-20% SDS-PAGE gel.
Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (ET1703-75) at 1/5,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature. -
Immunofluorescence analysis of frozen mouse brain tissue with Rabbit anti-Glutamate receptor 1 antibody (ET1612-10) at 1/200 dilution.
The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) for about 2 minutes in microwave oven. The tissues were blocked in 10% negative goat serum for 1 hour at room temperature, washed with PBS, and then probed with the primary antibody (ET1612-10, green) at 1/200 dilution overnight at 4 ℃, washed with PBS. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. Nuclei were counterstained with DAPI (blue). -
Immunofluorescence analysis of frozen mouse cerebellum tissue with Rabbit anti-PSD95 antibody (ET1602-20) at 1/500 dilution.
The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) for about 2 minutes in microwave oven. The tissues were blocked in 10% negative goat serum for 1 hour at room temperature, washed with PBS, and then probed with the primary antibody (ET1602-20, green) at 1/500 dilution overnight at 4 ℃, washed with PBS. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. Nuclei were counterstained with DAPI (blue). -
Immunofluorescence analysis of frozen mouse hippocampus tissue with Rabbit anti-NMDAR1 antibody (ET1703-75) at 1/200 dilution.
The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) for about 2 minutes in microwave oven. The tissues were blocked in 10% negative goat serum for 1 hour at room temperature, washed with PBS, and then probed with the primary antibody (ET1703-75, green) at 1/200 dilution overnight at 4 ℃, washed with PBS. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. Nuclei were counterstained with DAPI (blue).
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