Anti-HP1 gamma/CBX3 antibody [EPR19803]

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-HP1 gamma/CBX3 antibody [EPR19803] (AB213167)

Immunofluorescent analysis of 4% paraformaldehyde-fixed 0.1% Triton X-100 permeabilized HeLa (Human epithelial cell line from cervix adenocarcinoma) cells labeling HP1 gamma/CBX3 with ab213167 at 1/100 dilution followed by Goat Anti-Rabbit IgG (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

Confocal image showing nuclear staining on HeLa cells.

The nuclear counterstain is DAPI (blue).

Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) at 1/1000 dilution and Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed (ab150120) at 1/1000 dilution (red).

The negative controls are as follows : -

-ve control 1 : ab213167 at 1/100 dilution followed by ab150120 at 1/1000 dilution.

-ve control 2 : ab7291 at 1/1000 dilution followed by ab150077 at 1/1000 dilution.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HP1 gamma/CBX3 antibody [EPR19803] (AB213167)

Immunohistochemical analysis of paraffin-embedded Human colon tissue labeling HP1 gamma/CBX3 with ab213167 at 1/250 dilution followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

Nucleus staining on epithelial cells of Human colon is observed.

Counter stained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody secondary antibody is ab97051 at 1/500 dilution.

Perform heat mediated antigen retrieval with EDTA buffer pH 9 before commencing with IHC staining protocol.

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-HP1 gamma/CBX3 antibody [EPR19803] (AB213167)

Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed HeLa (Human epithelial cell line from cervix adenocarcinoma) cells labeling HP1 gamma/CBX3with ab213167 at 1/60 dilution (red) compared with a Rabbit IgG,monoclonal-Isotype control (ab172730) (black) and an unlabelled 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.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HP1 gamma/CBX3 antibody [EPR19803] (AB213167)

Immunohistochemical analysis of paraffin-embedded Human cervix carcinoma tissue labeling HP1 gamma/CBX3 with ab213167 at 1/250 dilution followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

Nucleus staining on cancer cells of Human cervix carcinoma is observed.

Counter stained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody secondary antibody is ab97051 at 1/500 dilution.

Perform heat mediated antigen retrieval with EDTA buffer pH 9 before commencing with IHC staining protocol.

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-HP1 gamma/CBX3 antibody [EPR19803] (AB213167)

Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed NIH/3T3 (Mouse embryonic fibroblast cell line) cells labeling HP1 gamma/CBX3 with ab213167 at 1/60 dilution (red) compared with a Rabbit IgG,monoclonal- Isotype control (ab172730) (black) and an unlabelled 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.

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-HP1 gamma/CBX3 antibody [EPR19803] (AB213167)

Immunofluorescent analysis of 4% paraformaldehyde-fixed 0.1% Triton X-100 permeabilized NIH/3T3 (Mouse embryonic fibroblast cell line) cells labeling HP1 gamma/CBX3 with ab213167 at 1/100 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 cells.

Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) at 1/1000 dilution and Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed (ab150120) at 1/1000 dilution (red).

The negative controls are as follows : -

-ve control 1 : ab213167 at 1/100 dilution followed by ab150120 at 1/1000 dilution.

-ve control 2 : ab7291 at 1/1000 dilution followed by ab150077 at 1/1000 dilution.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HP1 gamma/CBX3 antibody [EPR19803] (AB213167)

Immunohistochemical analysis of paraffin-embedded Mouse testis tissue labeling HP1 gamma/CBX3 with ab213167 at 1/250 dilution followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

Nucleus staining on Mouse testis is observed.

Counter stained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody secondary antibody is ab97051 at 1/500 dilution.

Perform heat mediated antigen retrieval with EDTA buffer pH 9 before commencing with IHC staining protocol.

• WB

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Western blot - Anti-HP1 gamma/CBX3 antibody [EPR19803] (AB213167)

Blocking/Dilution buffer : 5% NFDM/TBST.

Exposure time : Lane 1 : 3 minutes; Lane 2 : 15 seconds.

All lanes:

Western blot - Anti-HP1 gamma/CBX3 antibody [EPR19803] (ab213167) at 1/1000 dilution

Lane 1:

HepG2 (Human liver hepatocellular carcinoma cell line) whole cell lysate at 20 µg

Lane 2:

HeLa (Human epithelial cell line from cervix adenocarcinoma) 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: 20 kDa

Observed band size: 21 kDa

false

• WB

Supplier Data

Western blot - Anti-HP1 gamma/CBX3 antibody [EPR19803] (AB213167)

All lanes:

Western blot - Anti-HP1 gamma/CBX3 antibody [EPR19803] (ab213167) at 1/1000 dilution

Lane 1:

Human fetal brain lysate at 10 µg

Lane 2:

Human fetal kidney lysate at 10 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG Peroxidase Conjugate, specific to the non-reduced form of IgG at 1/10000 dilution

Predicted band size: 20 kDa

Observed band size: 21 kDa

false

Exposure time: 3min

• WB

Lab

Western blot - Anti-HP1 gamma/CBX3 antibody [EPR19803] (AB213167)

Lanes :
Lane 1 : Wild type HAP1 whole cell lysate (20 μg)
Lane 2 : CBX3 (KO) knockout HAP1 whole cell lysate (20 μg)
Lane 3 : HUVEC whole cell lysate (20 μg)
Lane 4 : HepG2 whole cell lysate (20 μg)

Lanes 1 - 4 : Merged signal (red and green). Green - ab213167 observed at 23 kDa. Red - loading control, ab8245, observed at 37 kDa.

ab213167 was shown to recognize HP1 gamma/CBX3 when HP1 gamma/CBX3 knockout samples were used, along with additional cross-reactive bands. Wild-type and HP1 gamma/CBX3 knockout samples were subjected to SDS-PAGE. ab213167 and ab8245 (Mouse anti GAPDH loading control) were incubated overnight at 4°C at 500 dilution and 1/10000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed ab216776 secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-HP1 gamma/CBX3 antibody [EPR19803] (ab213167)

Predicted band size: 20 kDa

false

• WB

Supplier Data

Western blot - Anti-HP1 gamma/CBX3 antibody [EPR19803] (AB213167)

Blocking/Dilution buffer : 5% NFDM/TBST.

Exposure time : Lane 1 and 2 : 3 minutes; Lane 3 : 10 seconds; Lane 4 : 30 seconds; Lane 5 : 10 seconds; Lane 6,7,8 and 9 : 30 seconds.

All lanes:

Western blot - Anti-HP1 gamma/CBX3 antibody [EPR19803] (ab213167) at 1/1000 dilution

Lane 1:

Mouse brain lysate at 10 µg

Lane 2:

Mouse kidney lysate at 10 µg

Lane 3:

Mouse spleen lysate at 10 µg

Lane 4:

Rat brain lysate at 10 µg

Lane 5:

Rat spleen lysate at 10 µg

Lane 6:

C6 (Rat glial tumor cell line) whole cell lysate at 10 µg

Lane 7:

RAW 264.7 (Mouse macrophage cell line transformed with Abelson murine leukemia virus) whole cell lysate at 10 µg

Lane 8:

PC-12 (Rat adrenal gland pheochromocytoma cell line) whole cell lysate at 10 µg

Lane 9:

NIH/3T3 (Mouse embryonic fibroblast cell line) whole cell lysate at 10 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: 20 kDa

Observed band size: 21 kDa

false