Anti-CHD1L antibody [EPR14515(2)]

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CHD1L antibody [EPR14515(2)] (AB197019)

Immunohistochemical analysis of paraffin-embedded Human hepatocellular carcinoma tissue labeling CHD1L with ab197019 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) secondary antibody at 1/500 dilution. Nuclear and weakly cytoplasm staining on Human hepatocellular carcinoma tissue is observed. Counter stained with Hematoxylin.

Negative control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

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

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-CHD1L antibody [EPR14515(2)] (AB197019)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cells from cervix adenocarcinoma) cells labeling CHD1L with ab197019 at 1/500 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/500 dilution (green). Nuclear and weakly cytoplasm staining on HeLa cell line was observed. The nuclear counter stain is DAPI (blue). Tubulin is detected with ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution and ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/500 dilution (red).
The negative controls are as follows :
-ve control 1 : ab197019 at 1/500 dilution followed by ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/500 dilution.
-ve control 2 : ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution followed by ab150077 (Alexa Fluor®488 Goat Anti-Rabbit IgG H&L) at 1/500 dilution.

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-CHD1L antibody [EPR14515(2)] (AB197019)

Intracellular flow cytometric analysis of HeLa (Human epithelial cells from cervix adenocarcinoma)cells labeling CHD1L with ab197019 at 1/520 dilution (red) compared with a rabbit monoclonal IgG isotype control (black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody; blue). Goat anti rabbit IgG (FITC) at 1/150 dilution was used as the secondary antibody.

• IP

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Immunoprecipitation - Anti-CHD1L antibody [EPR14515(2)] (AB197019)

CHD1L was immunoprecipitated from 293 (Human embryonic kidney) whole cell extract with ab197019 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab197019 at 1/1000 dilution. Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG, was used as secondary antibody at 1/1500 dilution. Lane 1 : 293 whole cell extract (Input) 10 μg. Lane 2 : ab197019 IP in 293 whole cell extract. Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab197019 in 293 whole cell extract.

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

All lanes:

Immunoprecipitation - Anti-CHD1L antibody [EPR14515(2)] (ab197019)

Predicted band size: 101 kDa

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• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CHD1L antibody [EPR14515(2)] (AB197019)

Immunohistochemical analysis of paraffin-embedded mouse liver tissue labeling CHD1L with ab197019 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) secondary antibody at 1/500 dilution. Nuclear and weakly cytoplasm staining on mouse liver tissue is observed. Counter stained with Hematoxylin.

Negative control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

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

• WB

Lab

Western blot - Anti-CHD1L antibody [EPR14515(2)] (AB197019)

Western blot : Rabbit Monoclonal [EPR14515(2)] to CHD1L ab197019 staining at 1/5000 dilution, shown in green; Mouse anti GAPDH (ab8245) loading control staining at 1/20,000 dilution, shown in magenta. A band was observed at 110 kDa in Wild-type U-87 MG cell lysates with no signal observed at this size in CHD1L knockout U-87 MG cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3pc 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-Mouse 800CW & Goat anti-Rabbit 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-CHD1L antibody [EPR14515(2)] (ab197019) at 1/5000 dilution

Lane 1:

Wild-type U-87 MG at 20 µg

Lane 2:

CHD1L knockout U-87 MG at 20 µg

Lane 3:

Saos-2 at 20 µg

Secondary

Lanes 1 - 3:

Goat anti-Mouse 800CW at 1/20000 dilution

Lanes 1 - 3:

Goat anti-Rabbit 680RD at 1/20000 dilution

Predicted band size: 101 kDa

Observed band size: 110 kDa,36 kDa

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• WB

Supplier Data

Western blot - Anti-CHD1L antibody [EPR14515(2)] (AB197019)

Blocking/Dilution buffer : 5% NFDM/TBST.

All lanes:

Western blot - Anti-CHD1L antibody [EPR14515(2)] (ab197019) at 1/5000 dilution

Lane 1:

HeLa (Human epithelial cells from cervix adenocarcinoma) cell lysate at 20 µg

Lane 2:

A549 (Human lung carcinoma) cell lysate at 20 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated at 1/1000 dilution

Predicted band size: 101 kDa

Observed band size: 101 kDa

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