All lanes : Anti-HDAC1 + HDAC2 antibody [EPR20327] (ab219054) at 1/10000 dilution

Lane 1 : His-tagged human HDAC2 recombinant protein (aa339-488)
Lane 2 : His-tagged human HDAC1 recombinant protein (aa1-482)

Lysates/proteins at 0.01 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

Predicted band size: 55 kDa
Observed band size: 22,60 kDa why is the actual band size different from the predicted?

Blocking/Dilution buffer: 5% NFDM/TBST.

Immunohistochemical analysis of paraffin-embedded human testis tissue labeling HDAC1 + HDAC2 with ab219054 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Nuclear staining on human testis is observed.

Counter stained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HEK-293 (Human epithelial cell line from embryonic kidney) cells labeling HDAC1 + HDAC2 with ab219054 at 1/1000 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

Confocal image showing nuclear staining on HEK-293 cell line.

The nuclear counterstain is DAPI (blue).

Tubulin is detected with ab195889 (Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594)) at 1/200 dilution (red).

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat anti-rabbit IgG (Alexa Fluor^® 488) (ab150077) at 1/1000 dilution.

All lanes : Anti-HDAC1 + HDAC2 antibody [EPR20327] (ab219054) at 1/2000 dilution

Lane 1 : HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate
Lane 2 : LNCaP (Human prostate cancer cell line) whole cell lysate
Lane 3 : SH-SY5Y (Human neuroblastoma cell line from bone marrow) whole cell lysate
Lane 4 : 293 (Human epithelial cell line from embryonic kidney) whole cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

Predicted band size: 55 kDa
Observed band size: 55 kDa


Exposure time: 5 seconds

Blocking/Dilution buffer: 5% NFDM/TBST.

All lanes : Anti-HDAC1 + HDAC2 antibody [EPR20327] (ab219054) at 1/1000 dilution

Lane 1 : Human fetal heart lysate
Lane 2 : Human fetal kidney lysate

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : VeriBlot for IP Detection Reagent (HRP) (ab131366) at 1/4000 dilution

Predicted band size: 55 kDa
Observed band size: 55 kDa


Exposure time: 3 minutes

Blocking/Dilution buffer: 5% NFDM/TBST.

All lanes : Anti-HDAC1 + HDAC2 antibody [EPR20327] (ab219054) at 1/1000 dilution

Lane 1 : Mouse brain lysate
Lane 2 : Mouse spleen lysate
Lane 3 : Rat brain lysate
Lane 4 : Rat kidney lysate
Lane 5 : Rat spleen lysate

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

Predicted band size: 55 kDa
Observed band size: 55 kDa

Blocking/Dilution buffer: 5% NFDM/TBST.

Exposure time: Lane 1/3-5: 3 minutes; Lane 2: 10 seconds.

All lanes : Anti-HDAC1 + HDAC2 antibody [EPR20327] (ab219054) at 1/1000 dilution

Lane 1 : C6 (Rat glial tumor cell line) whole cell lysate
Lane 2 : RAW 264.7 (Mouse macrophage cell line transformed with Abelson murine leukemia virus) whole cell lysate
Lane 3 : PC-12 (Rat adrenal gland pheochromocytoma cell line) whole cell lysate
Lane 4 : NIH/3T3 (Mouse embryonic fibroblast cell line) whole cell lysate

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

Predicted band size: 55 kDa
Observed band size: 55 kDa


Exposure time: 5 seconds

Blocking/Dilution buffer: 5% NFDM/TBST.

Immunohistochemical analysis of paraffin-embedded human tonsil tissue labeling HDAC1 + HDAC2 with ab219054 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Nuclear staining on lymphocytes of human tonsil is observed.

Counter stained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Immunohistochemical analysis of paraffin-embedded human prostate hyperplasia tissue labeling HDAC1 + HDAC2 with ab219054 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Nuclear staining on luminal epithelial cells of human prostate hyperplasia, but negative staining on basal cells.

Counter stained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Immunohistochemical analysis of paraffin-embedded human prostate cancer tissue labeling HDAC1 + HDAC2 with ab219054 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Nuclear staining on tumor cells of prostate cancer; weak or negative staining on basal cells.

Counter stained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Immunohistochemical analysis of paraffin-embedded human breast cancer tissue labeling HDAC1 + HDAC2 with ab219054 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Nuclear staining on tumor cells of human breast cancer is observed.

Counter stained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Immunohistochemical analysis of paraffin-embedded human synovial sarcoma tissue labeling HDAC1 + HDAC2 with ab219054 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Nuclear staining on human synovial sarcoma is observed.

Counter stained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Immunohistochemical analysis of paraffin-embedded mouse colon tissue labeling HDAC1 + HDAC2 with ab219054 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Nuclear staining on mouse colon is observed.

Counter stained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized NIH/3T3 (Mouse embryonic fibroblast cell line) cells labeling HDAC1 + HDAC2 with ab219054 at 1/1000 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

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

The nuclear counterstain is DAPI (blue).

Tubulin is detected with ab195889 (Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594)) at 1/200 dilution (red).

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat anti-rabbit IgG (Alexa Fluor^® 488) (ab150077) at 1/1000 dilution.

Flow cytometric analysis of 4% paraformaldehyde-fixed HeLa (Human epithelial cell line from cervix adenocarcinoma) cells labeling HDAC1 + HDAC2 with ab219054 at 1/700 dilution (red) compared with a rabbit monoclonal IgG isotype control (ab172730; black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody; blue). Goat anti rabbit IgG (Alexa Fluor^® 488) at 1/2000 dilution was used as the secondary antibody.

HDAC1 + HDAC2 was immunoprecipitated from 0.35 mg of HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate with ab219054 at 1/30 dilution.

Western blot was performed from the immunoprecipitate using ab219054 at 1/1000 dilution.

VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.

Lane 1: HeLa whole cell lysate 10 µg (Input).

Lane 2: ab219054 IP in HeLa whole cell lysate.

Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab219054 in HeLa whole cell lysate.

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

Exposure time: 1 second.