Anti-Ionotropic Glutamate receptor 2 antibody [EPR5032]

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Anti-Ionotropic Glutamate receptor 2 antibody [EPR5032] (AB133477)

Intracellular Flow Cytometry analysis of SH-SY5Y (Human neuroblastoma epithelial cell) cells labeling Ionotropic Glutamate receptor 2 (red) with ab133477 at a 1/200 dilution. Cells were fixed with 4% paraformaldehyde and permeabilized with 90% methanol. A goat anti-rabbit IgG (Alexa Fluorr^® 488) (ab150077) was used as the secondary antibody at a 1/2000 dilution. Black - Rabbit monoclonal IgG (Black) (ab172730). Blue (unlabeled control) - Cell without incubation with primary antibody and secondary antibody (Blue).

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-Ionotropic Glutamate receptor 2 antibody [EPR5032] (AB133477)

Immunocytochemistry analysis of Neuro-2a (Mouse neuroblastoma neuroblast) cells labeling Ionotropic Glutamate receptor 2 with purified ab133477 at 1/70 dilution (10 μg/mL). Cells were fixed in 4% Paraformaldehyde and permeabilized with 0.1% tritonX-100. Cells were counterstained with ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) 1/200 (2.5 μg/mL). Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) was used as the secondary antibody at 1/1000 (2 μg/mL) dilution. DAPI (blue) was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

• WB

Lab

Western blot - Anti-Ionotropic Glutamate receptor 2 antibody [EPR5032] (AB133477)

Blocking and diluting buffer and concentration : 5% NFDM/TBST

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/1000000 dilution.

Exposure time : ab133477 : 20s, ab181602 : 60s.

All lanes:

Western blot - Anti-Ionotropic Glutamate receptor 2 antibody [EPR5032] (ab133477) at 1/5000 dilution

Lane 1:

Mouse brain tissue whole cell lysate at 20 µg

Lane 2:

Rat brain tissue whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/ko/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Observed band size: 100 kDa

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• WB

Lab

Western blot - Anti-Ionotropic Glutamate receptor 2 antibody [EPR5032] (AB133477)

Blocking and diluting buffer and concentration : 5% NFDM/TBST

In Western blot, Anti-Myc tag antibody [9E10] - Loading Control (ab32) staining at 1/10000 dilution.

This antibody does not cross-react with Human GRIA1, GRIA3 and GRIA4.

Exposure time : ab133477 : 20s, ab32 : 80s.

All lanes:

Western blot - Anti-Ionotropic Glutamate receptor 2 antibody [EPR5032] (ab133477) at 1/5000 dilution

Lane 1:

293T transfected with an empty vector (vector control), containing a myc-His-tag®, whole cell lysate at 20 µg

Lane 2:

293T transfected with Human fragment GRIA2 expression vector containing a c-term myc-His-tag®, whole cell lysate at 20 µg

Lane 3:

293T transfected with Human full length GRIA3 expression vector containing a n-term myc-His-tag®, whole cell lysate at 20 µg

Lane 4:

293T transfected with human full length GRIA4 expression vector containing a c-term myc-His-tag®, whole cell lysate at 20 µg

Lane 5:

Recombinant Human full length GRIA1 Protein containing a C-Myc/DDK tag at 100 ng

Secondary

Lanes 1 - 5:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/ko/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Lanes 1 - 5:

Goat anti-mouse IgG (H+L), Peroxidase conjugated at 1/10000 dilution

Observed band size: 100 kDa

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• WB

Unknown

Western blot - Anti-Ionotropic Glutamate receptor 2 antibody [EPR5032] (AB133477)

All lanes:

Western blot - Anti-Ionotropic Glutamate receptor 2 antibody [EPR5032] (ab133477) at 1/5000 dilution

Lane 1:

Human hypothalamus lysate at 15 µg

Lane 2:

Human midbrain lysate at 15 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/2000 dilution

Predicted band size: 98 kDa

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• WB

CiteAb

Western blot - Anti-Ionotropic Glutamate receptor 2 antibody [EPR5032] (AB133477)

Western Blotting using Anti-Ionotropic Glutamate receptor 2 antibody [EPR5032], ab133477. Publication image from Zhou, Z. et al., 2019, Nat Commun, 31201320. Legend direct from paper.

R59X mice have an upregulation of NMDA receptors at the postsynaptic membrane. a Schematic of the R59X nonsense mutation at arginine 59, which results in early truncation of the catalytic domain of CDKL5. b Representative western blot results for several major ionotropic glutamate receptor subunits from postsynaptic density membrane fractions. Samples are from pairs of WT and R59X littermates. c–g R59X mice show a selective increase in level of GluN2B, a major subunit of the NMDA receptor (WT, n = 8 mice; R59X, n = 8 mice; unpaired t-test or Mann–Whitney test). Full-scan western blots of all samples are available in Supplemental Fig. 6. Source data are provided as a Source Data file

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• WB

CiteAb

Western blot - Anti-Ionotropic Glutamate receptor 2 antibody [EPR5032] (AB133477)

Western Blotting using Anti-Ionotropic Glutamate receptor 2 antibody [EPR5032], ab133477. Publication image from Zhou, Z. et al., 2019, Nat Commun, 31201320. Legend direct from paper.

Dlx-cKO mice have an upregulation of NMDA receptors at the postsynaptic membrane. a Representative western blot results for several major ionotropic glutamate receptor subunits from postsynaptic density membrane fractions. Samples are from pairs of WT and Dlx-cKO littermates. b–f Dlx-cKO mice show a selective increase in levels of GluN1 and GluN2B, two of the major subunits that compose the NMDA receptor (WT, n = 10 mice; Dlx-cKO, n = 10 mice; unpaired t-test or Mann–Whitney test). Full-scan western blots of all samples are available in Supplemental Fig. 6. *p < 0.05. Source data are provided as a Source Data file

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