Anti-HDAC1 antibody [EPR23847-170] - Nuclear Loading Control

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HDAC1 antibody [EPR23847-170] - Nuclear Loading Control (AB280198)

Immunohistochemical analysis of paraffin-embedded Human liver tissue labeling HDAC1 with ab280198 at 1/2000 dilution followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Nuclear staining on human liver (PMID : 18264140). The section was incubated with ab280198 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 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 [EPR23847-170] - Nuclear Loading Control (AB280198)

Immunohistochemical analysis of paraffin-embedded Human tonsil tissue labeling HDAC1 with ab280198 at 1/2000 dilution followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Nuclear staining on human tonsil (PMID : 23109994). The section was incubated with ab280198 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 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 [EPR23847-170] - Nuclear Loading Control (AB280198)

Flow cytometric analysis of 4% paraformaldehyde-fixe,d 90% methanol-permeabilized HeLa (Human cervix adenocarcinoma epithelial cell) cells labelling HDAC1 with ab280198 at 1/500 dilution (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.

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-HDAC1 antibody [EPR23847-170] - Nuclear Loading Control (AB280198)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa cells labelling HDAC1 with ab280198 at 1/5000 dilution, followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) antibody at 1/1000 dilution (Green). Confocal image showing nuclear and weakly cytoplasmic staining in HeLa cell line. 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).

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

• IP

Supplier Data

Immunoprecipitation - Anti-HDAC1 antibody [EPR23847-170] - Nuclear Loading Control (AB280198)

HDAC1 was immunoprecipitated from 0.35 mg HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate with ab280198 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab280198 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP)(ab131366) was used at 1/5000 dilution.

Lane 1 : HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate 10 ug

Lane 2 : ab280198 IP in HeLa whole cell lysate 10 ug

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab280198 in HeLa whole cell lysate

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

Exposure time : 1 second.

All lanes:

Immunoprecipitation - Anti-HDAC1 antibody [EPR23847-170] - Nuclear Loading Control (ab280198)

Predicted band size: 55 kDa

Observed band size: 62 kDa

false

• ChIP

Supplier Data

ChIP - Anti-HDAC1 antibody [EPR23847-170] - Nuclear Loading Control (AB280198)

Chromatin was prepared from K-562 cells according to the Abcam Dual-X-ChIP protocol*. Cells were fixed with 1.5 mM EGS for 30mins and then formaldehyde for 10min.
The ChIP was performed with 25 μg of chromatin, 5 μg of ab280198 (red), or 5 μg of rabbit normal IgG ab172730 (gray) and 25 μl of Protein A/G Dynabeads. The immunoprecipitated DNA was quantified by real time PCR (Sybr green approach).

*https : //www.abcam.com/resources?keywords=X%20ChIP%20protocol

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Anti-HDAC1 antibody [EPR23847-170] - Nuclear Loading Control (AB280198)

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized C6 (rat glial tumor glial cell) cells labelling HDAC1 with ab280198 at 1/500 dilution (0.1µg) followed by a Goat Anti-Rabbit IgG (Alexa Fluor^® 488, ab150081) at a 1/5000 dilution (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).

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HDAC1 antibody [EPR23847-170] - Nuclear Loading Control (AB280198)

Immunohistochemical analysis of paraffin-embedded Rat liver tissue labeling HDAC1 with ab280198 at 1/2000 dilution followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Nuclear staining on rat liver. The section was incubated with ab280198 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 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 [EPR23847-170] - Nuclear Loading Control (AB280198)

Flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol-permeabilized NIH/3T3 (Mouse embryonic fibroblast) cells labelling HDAC1 with ab280198 at 1/500 dilution (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.

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-HDAC1 antibody [EPR23847-170] - Nuclear Loading Control (AB280198)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized NIH/3T3 cells labelling HDAC1 with ab280198 at 1/5000 dilution, followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) antibody at 1/1000 dilution (Green). Confocal image showing nuclear and weakly cytoplasmic staining in NIH/3T3 cell line. 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).

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

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HDAC1 antibody [EPR23847-170] - Nuclear Loading Control (AB280198)

Immunohistochemical analysis of paraffin-embedded Mouse liver tissue labeling HDAC1 with ab280198 at 1/2000 dilution followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Nuclear staining on mouse liver. The section was incubated with ab280198 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 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.

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-HDAC1 antibody [EPR23847-170] - Nuclear Loading Control (AB280198)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized C6 (rat glial tumor glial cell) cells labelling HDAC1 with ab280198 at 1/500 dilution followed by a ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed at a 1/1000 (2 µg) dilution (Green).

Confocal image showing nuclear staining in C6 cell line (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 (2.5µg/ml) dilution (Magenta). The Nuclear counterstain was DAPI (Blue).

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

• IP

Lab

Immunoprecipitation - Anti-HDAC1 antibody [EPR23847-170] - Nuclear Loading Control (AB280198)

HDAC1 was immunoprecipitated from C6 (rat glial tumor glial cell) whole cell lysate with ab280198 at 1/30 dilution (2µg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab280198 at 1/1000 dilution.

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

All lanes:

Immunoprecipitation - Anti-HDAC1 antibody [EPR23847-170] - Nuclear Loading Control (ab280198) at 1/1000 dilution

Lane 1:

C6 (rat glial tumor glial cell) whole cell lysate (Input) at 10 µg

Lane 2:

C6 (rat glial tumor glial cell) whole cell lysate (+)

Lane 3:

Rabbit monoclonal IgG (<a href='/en-us/products/primary-antibodies/rabbit-igg-monoclonal-epr25a-isotype-control-ab172730'>ab172730</a>) instead of ab280198 in C6 whole cell lysate (-)

Secondary

All lanes:

Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/5000 dilution

Observed band size: 55 kDa

false

Exposure time: 8s

• IP

Supplier Data

Immunoprecipitation - Anti-HDAC1 antibody [EPR23847-170] - Nuclear Loading Control (AB280198)

HDAC1 was immunoprecipitated from 0.35 mg NIH/3T3 (mouse embryonic fibroblast) whole cell lysate with ab280198 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab280198 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP)(ab131366) was used at 1/5000 dilution.

Lane 1 : NIH/3T3 (mouse embryonic fibroblast) whole cell lysate 10 ug

Lane 2 : ab280198 IP in NIH/3T3 whole cell lysate 10 ug

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab280198 in NIH/3T3 whole cell lysate

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

Exposure time : 1 second.

All lanes:

Immunoprecipitation - Anti-HDAC1 antibody [EPR23847-170] - Nuclear Loading Control (ab280198)

Predicted band size: 55 kDa

Observed band size: 62 kDa

false

• WB

Lab

Western blot - Anti-HDAC1 antibody [EPR23847-170] - Nuclear Loading Control (AB280198)

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

The observed MW is consistent with what has been described in the literature (PMID : 24551070).

Exposure time : 37 seconds.

All lanes:

Western blot - Anti-HDAC1 antibody [EPR23847-170] - Nuclear Loading Control (ab280198) at 1/5000 dilution

Lane 1:

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

Lane 2:

K-562 (human chronic myelogenous leukemia lymphoblast) whole cell lysate at 20 µg

Lane 3:

HeLa (human cervix adenocarcinoma epithelial 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 1/100000 dilution

Predicted band size: 55 kDa

Observed band size: 62 kDa

false

• WB

Lab

Western blot - Anti-HDAC1 antibody [EPR23847-170] - Nuclear Loading Control (AB280198)

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

The observed MW is consistent with what has been described in the literature (PMID : 24551070).

Exposure time : 15 seconds.

All lanes:

Western blot - Anti-HDAC1 antibody [EPR23847-170] - Nuclear Loading Control (ab280198) at 1/1000 dilution

Lane 1:

Human heart tissue lysate at 20 µg

Lane 2:

Human kidney tissue lysate at 20 µg

Secondary

All lanes:

Western blot - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/1000 dilution

Predicted band size: 55 kDa

Observed band size: 62 kDa

false

• WB

Supplier Data

Western blot - Anti-HDAC1 antibody [EPR23847-170] - Nuclear Loading Control (AB280198)

ab280198 Anti-HDAC1 antibody [EPR23847-170] was shown to specifically react with HDAC1 in wild-type HAP1 cells. Loss of signal was observed when knockout cell line (knockout cell lysate) was used. Wild-type and HDAC1 knockout samples were subjected to SDS-PAGE. ab280198 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated at room temperature for 2. 5 hours at 1/1000 dilution and 1/20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-HDAC1 antibody [EPR23847-170] - Nuclear Loading Control (ab280198) at 1/1000 dilution

Lane 1:

Wild-type HAP1 cell lysate at 20 µg

Lane 2:

HDAC1 knockout HAP1 cell lysate at 40 µg

Lane 3:

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

Lane 4:

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

Secondary

All lanes:

Goat Anti-Rabbit IgG H&L (IRDye® 800CW) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) and Goat Anti-Mouse IgG H&L (IRDye® 680RD) (<a href='/en-us/products/secondary-antibodies/goat-mouse-igg-h-l-irdye-680rd-preadsorbed-ab216776'>ab216776</a>) at 1/10000 dilution

Predicted band size: 55 kDa

false

• ChIC/CUT&RUN-seq

Supplier Data

ChIC/CUT&RUN sequencing - Anti-HDAC1 antibody [EPR23847-170] - Nuclear Loading Control (AB280198)

ChIC/CUT&RUN was performed using a pAG-MNAse at a final concentration of 700 ng/mL, 2 x 10^5 K-562 (Human chronic myelogenous leukemia lymphoblast) cells and 5 µg of ab280198 [EPR23847-170]. The resulting DNA was sequenced on the Illumina NovaSeq 6000 to a depth of 10 million reads. The negative IgG control ab172730 is also shown. Additional screenshots of mapped reads can be found in the Protocol booklet in the Product Protocol section. The University of Geneva owns patents relevant to ChIC (Chromatin Immuno-Cleavage) methods.

• WB

Lab

Western blot - Anti-HDAC1 antibody [EPR23847-170] - Nuclear Loading Control (AB280198)

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

The observed MW is consistent with what has been described in the literature (PMID : 24551070).

Exposure time : 37 seconds.

All lanes:

Western blot - Anti-HDAC1 antibody [EPR23847-170] - Nuclear Loading Control (ab280198) at 1/5000 dilution

Lane 1:

C6 (rat glial tumor glial cell) whole cell lysate at 20 µg

Lane 2:

RAW264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysate at 20 µg

Lane 3:

PC-12 (rat adrenal gland pheochromocytoma) whole cell lysate at 20 µg

Lane 4:

NIH/3T3 (mouse embryonic fibroblast) 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 1/100000 dilution

Predicted band size: 55 kDa

Observed band size: 62 kDa

false

• WB

Lab

Western blot - Anti-HDAC1 antibody [EPR23847-170] - Nuclear Loading Control (AB280198)

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

This antibody has no cross-reaction with human HDAC2.

These recombinant proteins were made in house. These two recombinant proteins were expressed from E.coli expression systems.

Exposure time : 10 seconds.

All lanes:

Western blot - Anti-HDAC1 antibody [EPR23847-170] - Nuclear Loading Control (ab280198) at 1/1000 dilution

Lane 1:

His-tagged human HDAC1 recombinant protein (aa1-482) at 0.01 µg

Lane 2:

His-tagged human HDAC2 recombinant protein (aa1-488) at 0.01 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 1/100000 dilution

Predicted band size: 55 kDa

Observed band size: 66 kDa

false