Anti-HDAC1 antibody [RM1004] - BSA and Azide free

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-HDAC1 antibody [RM1004] - BSA and Azide free (AB283864)

This data was developed using ab281585, the same antibody clone in a different buffer formulation.Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa cells labelling HDAC1 with ab281585 at 1/50 (10.4 µg/ml) dilution, followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) antibody at 1/1000 dilution (Green). ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor^® 594) was used to counterstain tubulin at 1/200 dilution (Red). The nuclear counterstain was DAPI (Blue).

Confocal image showing mostly nuclear staining in HeLa cell line.

Secondary antibody only control : Secondary antibody is ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) at 1/1000 dilution.

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-HDAC1 antibody [RM1004] - BSA and Azide free (AB283864)

This data was developed using ab281585, the same antibody clone in a different buffer formulation.Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized HeLa (Human cervix adenocarcinoma cell) cells labelling HDAC1 with ab281585 at 1/500 dilution (0.1 µg) (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). A Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) at 1/2000 dilution was used as the secondary antibody.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HDAC1 antibody [RM1004] - BSA and Azide free (AB283864)

This data was developed using ab281585, the same antibody clone in a different buffer formulation.Immunohistochemical analysis of paraffin-embedded human testis tissue labelling HDAC1 with ab281585 at 1/4000 (0.13 µg/ml) dilution followed by ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

Nuclear staining on human testis (PMID : 16960727).

The section was incubated with ab281585 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 ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.

• IP

Supplier Data

Immunoprecipitation - Anti-HDAC1 antibody [RM1004] - BSA and Azide free (AB283864)

This data was developed using ab281585, the same antibody clone in a different buffer formulation.HDAC1 was immunoprecipitated from 0.35 mg Jurkat (human T cell leukemia T lymphocyte), whole cell lysate 10 µg with ab281585 at 1/30 dilution (2 µg in 0.35 mg lysates). Western blot was performed on the immunoprecipitate using ab281585 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP)(ab131366) was used at 1/5000 dilution.

Lane 1 : Jurkat whole cell lysate 10 µg

Lane 2 : ab281585 IP in Jurkat whole cell lysate

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab281585 in Jurkat whole cell lysate

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

Exposure time : 3 seconds.

All lanes:

Immunoprecipitation - Anti-HDAC1 antibody [RM1004] - Nuclear Loading Control (<a href='/en-us/products/primary-antibodies/hdac1-antibody-rm1004-nuclear-loading-control-ab281585'>ab281585</a>)

Predicted band size: 55 kDa

false

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HDAC1 antibody [RM1004] - BSA and Azide free (AB283864)

This data was developed using ab281585, the same antibody clone in a different buffer formulation.Immunohistochemical analysis of paraffin-embedded mouse testis tissue labelling HDAC1 with ab281585 at 1/4000 (0.13 µg/ml) dilution followed by ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

Nuclear staining on mouse testis.

The section was incubated with ab281585 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 ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

Heat mediated antigen retrieval 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-HDAC1 antibody [RM1004] - BSA and Azide free (AB283864)

This data was developed using ab281585, the same antibody clone in a different buffer formulation.Immunohistochemical analysis of paraffin-embedded rat testis tissue labelling HDAC1 with ab281585 at 1/4000 (0.13 µg/ml) dilution followed by ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

Nuclear staining on rat testis.

The section was incubated with ab281585 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 ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-HDAC1 antibody [RM1004] - BSA and Azide free (AB283864)

This data was developed using ab281585, the same antibody clone in a different buffer formulation.Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized NIH/3T3 (Mouse embryonic fibroblast) cells labelling HDAC1 with ab281585 at 1/500 dilution (0.1 µg) (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). A Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) at 1/2000 dilution was used as the secondary antibody.

• IHC-Fr

Supplier Data

Immunohistochemistry (Frozen sections) - Anti-HDAC1 antibody [RM1004] - BSA and Azide free (AB283864)

This data was developed using ab281585, the same antibody clone in a different buffer formulation.Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen rat hippocampus tissue labeling HDAC1 with ab281585 at 1/100 (5.55 µg/ml) dilution followed by ab150077 AlexaFluor^®488 Goat anti-Rabbit secondary at 1/1000 dilution (Green). The nuclear counterstain was DAPI (Blue).

Nuclear staining on rat hippocampus is observed.

Secondary antibody control : Secondary antibody is ab150077 AlexaFluor^®488 Goat anti-Rabbit secondary at 1/1000 dilution.

Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20).

• IHC-Fr

Supplier Data

Immunohistochemistry (Frozen sections) - Anti-HDAC1 antibody [RM1004] - BSA and Azide free (AB283864)

This data was developed using ab281585, the same antibody clone in a different buffer formulation.Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen mouse hippocampus tissue labeling HDAC1 with ab281585 at 1/100 (5.55 µg/ml) dilution followed by ab150077 AlexaFluor^®488 Goat anti-Rabbit secondary at 1/1000 dilution (Green). The nuclear counterstain was DAPI (Blue).

Nuclear staining on mouse hippocampus is observed.

Secondary antibody control : Secondary antibody is ab150077 AlexaFluor^®488 Goat anti-Rabbit secondary at 1/1000 dilution.

Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20).

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-HDAC1 antibody [RM1004] - BSA and Azide free (AB283864)

This data was developed using ab281585, the same antibody clone in a different buffer formulation.Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized NIH/3T3 cells labelling HDAC1 with ab281585 at 1/50 (10.4 µg/ml) dilution, followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) antibody at 1/1000 dilution (Green). ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor^® 594) was used to counterstain tubulin at 1/200 dilution (Red). The nuclear counterstain was DAPI (Blue).

Confocal image showing mostly nuclear staining in NIH/3T3 cell line.

Secondary antibody only control : Secondary antibody is ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) at 1/1000 dilution.

• WB

Lab

Western blot - Anti-HDAC1 antibody [RM1004] - BSA and Azide free (AB283864)

This data was developed using ab281585, the same antibody clone in a different buffer formulation.

False colour image of Western blot : Anti-HDAC1 antibody [RM1004] staining at 1/1000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab281585 was shown to bind specifically to HDAC1. A band was observed at 60 kDa in wild-type HAP1 cell lysates with no signal observed at this size in HDAC1 knockout cell line. To generate this image, wild-type and HDAC1 knockout HAP1 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 5 % milk in TBS-0.1 % Tween^® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) at 1/20000 dilution.

All lanes:

Western blot - Anti-HDAC1 antibody [RM1004] - BSA and Azide free (ab283864) at 1/1000 dilution

Lane 1:

Wild-type HAP1 cell lysate at 40 µg

Lane 2:

HDAC1 knockout HAP1 cell lysate at 40 µg

Predicted band size: 55 kDa

Observed band size: 60 kDa

false

• WB

Lab

Western blot - Anti-HDAC1 antibody [RM1004] - BSA and Azide free (AB283864)

This data was developed using ab281585, the same antibody clone in a different buffer formulation.

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

Exposure time : 5 seconds

All lanes:

Western blot - Anti-HDAC1 antibody [RM1004] - Nuclear Loading Control (<a href='/en-us/products/primary-antibodies/hdac1-antibody-rm1004-nuclear-loading-control-ab281585'>ab281585</a>) at 1/1000 dilution

Lane 1:

HeLa (human epithelial cell line from cervix adenocarcinoma), whole cell lysate at 20 µg

Lane 2:

Jurkat (human T cell leukemia T lymphocyte), whole cell lysate at 20 µg

Lane 3:

NIH/3T3 (mouse embryonic fibroblast), whole cell lysate at 20 µg

Lane 4:

C6 (rat glial tumor glial cell), whole cell 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 1/50000 dilution

Predicted band size: 55 kDa

false