Anti-HuD antibody [EPR29580-516]

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HuD antibody [EPR29580-516] (AB324613)

Immunohistochemical analysis of paraffin-embedded Mouse liver tissue labeling HuD with ab324613 at 1/1000 (0.507 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Negative control : no staining on mouse liver. 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-HuD antibody [EPR29580-516] (AB324613)

Immunohistochemical analysis of paraffin-embedded Mouse spleen tissue labeling HuD with ab324613 at 1/1000 (0.507 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Negative control : no staining on mouse 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

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HuD antibody [EPR29580-516] (AB324613)

Immunohistochemical analysis of paraffin-embedded Rat spleen tissue labeling HuD with ab324613 at 1/1000 (0.507 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Negative control : no staining on rat 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

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HuD antibody [EPR29580-516] (AB324613)

Immunohistochemical analysis of paraffin-embedded Rat liver tissue labeling HuD with ab324613 at 1/1000 (0.507 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Negative control : no staining on rat liver. 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-Fr

Supplier Data

Immunohistochemistry (Frozen sections) - Anti-HuD antibody [EPR29580-516] (AB324613)

Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen Mouse liver (fresh frozen) tissue labeling HuD with ab324613 at 1/50 (10.14 ug/ml) dilution followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 (2 ug/mL) dilution (Green).

Negative control : confocal image showing no staining on mouse liver. The nuclear counterstain was DAPI (Blue). The section was incubated with ab324613 for 60 mins at room temperature. The section was then mounted using Fluoromount®.The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

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

• IHC-Fr

Supplier Data

Immunohistochemistry (Frozen sections) - Anti-HuD antibody [EPR29580-516] (AB324613)

Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen Rat liver (fresh frozen) tissue labeling HuD with ab324613 at 1/50 (10.14 ug/ml) dilution followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 (2 ug/mL) dilution (Green).

Negative control : confocal image showing no staining on rat liver. The nuclear counterstain was DAPI (Blue). The section was incubated with ab324613 for 60 mins at room temperature. The section was then mounted using Fluoromount®.The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

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

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-HuD antibody [EPR29580-516] (AB324613)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized mouse splenocytes labelling HuD with ab324613 at 1/50 (10.14 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 (2ug/ml) dilution (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).

Anti-mouse CD3 rat monoclonal antibody (Alexa Fluor® 647) was used to counterstain tubulin at 1/100 dilution (Magenta).

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

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-HuD antibody [EPR29580-516] (AB324613)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized rat splenocytes labelling HuD with ab324613 at 1/50 (10.14 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 (2ug/ml) dilution (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).

Anti-rat CD3 mouse monoclonal antibody (Alexa Fluor® 647) was used to counterstain tubulin at 1/100 5ug/ml dilution (Magenta).

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

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HuD antibody [EPR29580-516] (AB324613)

Immunohistochemical analysis of paraffin-embedded Rat colon tissue labeling HuD with ab324613 at 1/1000 (0.507 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on myenteric nerve plexus of rat colon. 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-HuD antibody [EPR29580-516] (AB324613)

Immunohistochemical analysis of paraffin-embedded Mouse cerebrum tissue labeling HuD with ab324613 at 1/1000 (0.507 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on neurons of mouse cerebrum (PMID : 20584986; PMID : 22387028). 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-Fr

Supplier Data

Immunohistochemistry (Frozen sections) - Anti-HuD antibody [EPR29580-516] (AB324613)

Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen Rat cerebrum (fresh frozen) tissue labeling HuD with ab324613 at 1/50 (10.14 ug/ml) dilution followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 (2 ug/mL) dilution (Green).

Panel A : merged staining of anti-ELAVL4 (ab324613, green), anti-NeuN (ab190565, magenta) on rat cerebrum.
Panel B : anti-ELAVL4 stained on rat cerebrum.
Panel C : anti-NeuN stained in neurons of rat cerebrum.
The section was incubated in two rounds of staining : in the order of ab324613 and ab190565, for 1 hr at room temperature. The nuclear counterstain was DAPI (Blue). The section was then mounted using Fluoromount®.The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HuD antibody [EPR29580-516] (AB324613)

Immunohistochemical analysis of paraffin-embedded Mouse colon tissue labeling HuD with ab324613 at 1/1000 (0.507 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on myenteric nerve plexus of mouse colon. 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-HuD antibody [EPR29580-516] (AB324613)

Immunohistochemical analysis of paraffin-embedded Rat cerebrum tissue labeling HuD with ab324613 at 1/1000 (0.507 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on neurons of rat cerebrum. 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-Fr

Supplier Data

Immunohistochemistry (Frozen sections) - Anti-HuD antibody [EPR29580-516] (AB324613)

Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen Mouse cerebrum (fresh frozen) tissue labeling HuD with ab324613 at 1/50 (10.14 ug/ml) dilution followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 (2 ug/mL) dilution (Green).

Panel A : merged staining of anti-ELAVL4 (ab324613, green), anti-NeuN (ab190565, magenta) on mouse cerebrum.
Panel B : anti-ELAVL4 stained on mouse cerebrum.
Panel C : anti-NeuN stained in neurons of mouse cerebrum.
The section was incubated in two rounds of staining : in the order of ab324613 and ab190565, for 1 hr at room temperature. The nuclear counterstain was DAPI (Blue). The section was then mounted using Fluoromount®.The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

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

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-HuD antibody [EPR29580-516] (AB324613)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized mouse primary neurons labelling HuD with ab324613 at 1/50 (10.14 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 (2ug/ml) dilution (Green).

Confocal image showing cytoplasmic and nuclear staining in mouse primary neuron (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).

ab11267 Anti-MAP2 mouse monoclonal antibody was used to counterstain tubulin at 1/500 (4ug/ml) dilution, followed by ab150120 Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) at 1/1000 (2ug/ml) dilution (Magenta).

-ve control 1 : ab324613 at a 1/50 dilution followed by ab150120 at a 1/1000 dilution.

-ve control 2 : ab11267 at a 1/500 dilution followed by ab150081 at a 1/1000 dilution.

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-HuD antibody [EPR29580-516] (AB324613)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized rat primary neurons labelling HuD with ab324613 at 1/50 (10.14 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 (2ug/ml) dilution (Green).

Confocal image showing cytoplasmic and nuclear staining in rat primary neuron (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).

ab11267 Anti-MAP2 mouse monoclonal antibody was used to counterstain tubulin at 1/500 (4ug/ml) dilution, followed by ab150120 Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) at 1/1000 (2ug/ml) dilution (Magenta).

-ve control 1 : ab324613 at a 1/50 dilution followed by ab150120 at a 1/1000 dilution.

-ve control 2 : ab11267 at a 1/500 dilution followed by ab150081 at a 1/1000 dilution.

• I-ELISA

Supplier Data

Indirect ELISA - Anti-HuD antibody [EPR29580-516] (AB324613)

Indirect ELISA analysis of ab324613 at 1000-0 ng/ml. The Secondary antibody used was Peroxidase AffiniPure Goat Anti-Rabbit IgG, Fc fragment specific at 1 : 20000 dilution dilution.

Antigen : Mouse HuR/ELAVL1, Mouse HuB/ELAVL2, Mouse HuC/ELAVL3, Mouse HuD/ELAVL4.

Antigen concentration : 1000 ng/ml

This antibody does not cross-react with Mouse HuR/ELAVL1, Mouse HuB/ELAVL2, Mouse HuC/ELAVL3

• WB

Supplier Data

Western blot - Anti-HuD antibody [EPR29580-516] (AB324613)

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

Low expression : 4T1, A20

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

All lanes:

Western blot - Anti-HuD antibody [EPR29580-516] (ab324613) at 1/1000 dilution

Lane 1:

Beta-TC-6 (mouse pancreas insulinoma beta cell) whole cell lysate at 48 µg

Lane 2:

Neuro-2a (mouse neuroblastoma neuroblast) whole cell lysate at 48 µg

Lane 3:

4T1 (mouse mammary gland carcinoma epithelial cell) whole cell lysate at 48 µg

Lane 4:

A20 (mouse reticulum sarcoma B lymphocyte) whole cell lysate at 48 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 42 kDa,36 kDa

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-HuD antibody [EPR29580-516] (AB324613)

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

Negative control : kidney, spleen (PMID : 22387028; PMID : 20584986)

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

All lanes:

Western blot - Anti-HuD antibody [EPR29580-516] (ab324613) at 1/1000 dilution

Lane 1:

Rat cerebral cortex tissue lysate at 20 µg

Lane 2:

Rat brain tissue lysate at 20 µg

Lane 3:

Rat kidney tissue lysate at 20 µg

Lane 4:

Rat spleen tissue lysate at 20 µg

Lane 5:

Mouse cerebral cortex tissue lysate at 20 µg

Lane 6:

Mouse brain tissue lysate at 20 µg

Lane 7:

Mouse kidney tissue lysate at 20 µg

Lane 8:

Mouse spleen tissue lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 42 kDa,36 kDa

false

Exposure time: 180s