Anti-EAAT2 antibody [EPR29552-528]

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-EAAT2 antibody [EPR29552-528] (AB323867)

Immunohistochemical analysis of paraffin-embedded human astrocytoma tissue labeling EAAT2 with ab323867 at 1/2000 (0.263 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on human astrocytoma. The primary antibody was incubated for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-EAAT2 antibody [EPR29552-528] (AB323867)

Immunohistochemical analysis of paraffin-embedded human cardiac muscle tissue labeling EAAT2 with ab323867 at 1/2000 (0.263 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Negative control : no staining on human cardiac muscle. The primary antibody was incubated for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-EAAT2 antibody [EPR29552-528] (AB323867)

Immunohistochemical analysis of paraffin-embedded human cerebrum tissue labeling EAAT2 with ab323867 at 1/2000 (0.263 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on human cerebrum (PMID : 25391854). The primary antibody was incubated for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-EAAT2 antibody [EPR29552-528] (AB323867)

Immunohistochemical analysis of paraffin-embedded human spleen tissue labeling EAAT2 with ab323867 at 1/2000 (0.263 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Negative control : no staining on human spleen. The primary antibody was incubated for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-EAAT2 antibody [EPR29552-528] (AB323867)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized mouse primary neural/glia cells labelling EAAT2 with ab323867 at 1/100 dilution (5.26 ug/ml), followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 dilution (2 ug/ml) (Green).

Confocal image showing positive staining in mouse primary glia cells (shown in green). The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

Anti-GFAP mouse monoclonal antibody was used to counterstain tubulin at 1/50 dilution (10 ug/ml), followed by ab150120 Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) at 1/1000 dilution (2 ug/ml) (Magenta).

-ve control 1 : ab323867 at a 1/100 dilution followed by ab150120 at a 1/1000 dilution.

-ve control 2 : Anti-GFAP mouse monoclonal antibody at a 1/50 dilution followed by ab150081 at a 1/1000 dilution.

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-EAAT2 antibody [EPR29552-528] (AB323867)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized rat primary neural/glia cells labelling EAAT2 with ab323867 at 1/100 dilution (5.26 ug/ml), followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 dilution (2 ug/ml) (Green).

Confocal image showing positive staining in rat primary glia cells (shown in green). The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

ab10062 Anti-GFAP mouse monoclonal antibody was used to counterstain tubulin at 1/50 dilution (10 ug/ml), followed by ab150120 Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) at 1/1000 dilution (2 ug/ml) (Magenta).

-ve control 1 : ab323867 at a 1/100 dilution followed by ab150120 at a 1/1000 dilution.

-ve control 2 : ab10062 at a 1/50 dilution followed by ab150081 at a 1/1000 dilution.

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-EAAT2 antibody [EPR29552-528] (AB323867)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized mouse splenocytes labelling EAAT2 with ab323867 at 1/100 dilution (5.26 ug/ml), followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 dilution (2 ug/ml) (Green).

Negative control : Confocal image showing no staining in mouse splenocytes (shown in green). The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 dilution (2.5 ug/ml) (Magenta).

Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution (2 ug/ml).

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-EAAT2 antibody [EPR29552-528] (AB323867)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized rat splenocytes labelling EAAT2 with ab323867 at 1/100 dilution (5.26 ug/ml), followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 dilution (2 ug/ml) (Green).

Negative control : Confocal image showing no staining in rat splenocytes (shown in green). The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 dilution (2.5 ug/ml) (Magenta).

Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution (2 ug/ml).

• WB

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Western blot - Anti-EAAT2 antibody [EPR29552-528] (AB323867)

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

Negative control : spleen, testis (PMID : 1448170; PMID : 7698742).

The molecular weight observed is consistent with what has been described in the literature (PMID : 20193040; PMID : 15483603; PMID : 26483543).

Bands above 250KD are likely to be multimer forms of EAAT2 (PMID : 20193040).

Samples are non-boiled as boiling may cause protein aggregation.

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

All lanes:

Western blot - Anti-EAAT2 antibody [EPR29552-528] (ab323867) at 1/1000 dilution

Lane 1:

Human striatum tissue lysate at 20 µg

Lane 2:

Human hypothalamus tissue lysate at 20 µg

Lane 3:

Human spleen tissue lysate at 20 µg

Lane 4:

Human testis tissue lysate at 20 µg

Secondary

All lanes:

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

Observed band size: 65 kDa,180 kDa,36 kDa

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-EAAT2 antibody [EPR29552-528] (AB323867)

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

Negative control : spleen, testis (PMID : 1448170; PMID : 7698742).

The molecular weight observed is consistent with what has been described in the literature (PMID : 20193040; PMID : 15483603; PMID : 26483543).

Bands above 250KD are likely to be multimer forms of EAAT2 (PMID : 20193040).

Samples are non-boiled as boiling may cause protein aggregation.

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

All lanes:

Western blot - Anti-EAAT2 antibody [EPR29552-528] (ab323867) at 1/1000 dilution

Lane 1:

Mouse striatum tissue lysate at 20 µg

Lane 2:

Mouse hippocampus tissue lysate at 20 µg

Lane 3:

Mouse brain tissue lysate at 20 µg

Lane 4:

Mouse spleen tissue lysate at 20 µg

Lane 5:

Mouse testis tissue lysate at 20 µg

Lane 6:

Rat striatum tissue lysate at 20 µg

Lane 7:

Rat brain tissue lysate at 20 µg

Lane 8:

Rat spleen tissue lysate at 20 µg

Lane 9:

Rat testis tissue lysate at 20 µg

Secondary

All lanes:

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

Observed band size: 65 kDa,180 kDa,36 kDa

false

Exposure time: 3s

• Dot

Supplier Data

Dot Blot - Anti-EAAT2 antibody [EPR29552-528] (AB323867)

Dot blot analysis of EAAT2 using ab323867 at 1 : 1000 (0.526 ug/ml) followed by a Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) at 1 : 100,000 dilution.

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

This antibody does not cross-react with human EAAT1 and EAAT3.

In Dot Blot, Anti-6X His tag® antibody [EPR20547] - ChIP Grade (ab213204) staining at 1/5000 dilution.

All lanes:

Dot Blot - Anti-EAAT2 antibody [EPR29552-528] (ab323867) at 1/1000 dilution

Lane 1:

293T cells transfected with a human EAAT2 expression vector containing a myc-His-tag®, whole cell lysate

Lane 2:

293T cells transfected with a human EAAT1 expression vector containing a myc-His-tag®, whole cell lysate

Lane 3:

293T cells transfected with a human EAAT3 expression vector containing a myc-His-tag®, whole cell lysate

Secondary

All lanes:

Dot 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

false

Exposure time: 180s