Anti-IRAK4 antibody [Y279]

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-IRAK4 antibody [Y279] (AB32511)

Immunocytochemistry/Immunofluorescence analysis of Jurkat (Human T cell leukemia cell line from peripheral blood) cells labeling IRAK4 with ab32511 (unpurified) at 1/100 dilution. Cells were fixed with 100% methanol. ab150077, an AlexaFluor® 488 conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody. The cells were co-stained with ab195889, anti-alpha tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) at 1/200 dilution. Nuclei counterstained with DAPI (blue).

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-IRAK4 antibody [Y279] (AB32511)

Intracellular Flow Cytometry analysis of Jurkat (human acute T cell leukemia) cells labeling IRAK4 with purified ab32511 at 1/110 dilution (10ug/ml) (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. A Goat anti rabbit IgG (Alexa Fluor^® 488) (1/2000 dilution) was used as the secondary antibody. Rabbit monoclonal IgG (Black) was used as the isotype control, cells without incubation with primary antibody and secondary antibody (Blue) was used as the unlabeled control.

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-IRAK4 antibody [Y279] (AB32511)

Immunocytochemistry/ Immunofluorescence analysis of Jurkat (Human T cell leukemia T lymphocyte) cells labeling IRAK4 with purified ab32511 at 1 : 500 dilution (2.6 µg/ml). Cells were fixed in 100% Methanol and permeabilized with None. Cells were counterstained with none. Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) was used as the secondary antibody at 1 : 1000 (2 µg/ml) dilution. DAPI (blue) was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

• WB

Lab

Western blot - Anti-IRAK4 antibody [Y279] (AB32511)

All lanes:

Western blot - Anti-IRAK4 antibody [Y279] (ab32511) at 1/1000 dilution

All lanes:

K-562 (Human chronic myelogenous leukemia lymphoblast) whole cell lysates at 15 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 1/20000 dilution

Predicted band size: 52 kDa

Observed band size: 52 kDa

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• WB

Lab

Western blot - Anti-IRAK4 antibody [Y279] (AB32511)

Western blot : Rabbit Monoclonal [Y279] to IRAK4 ab32511 staining at 1/1000 dilution, shown in black. A band was observed at 55 kDa in MCF7 Pulldown with ab32511, enriched from the MCF7 input sample and no signal observed at this size in MCF7 Pulldown with Isotype Control ab172730. 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-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20,000 dilution.

Lanes 1 - 3:

Western blot - Anti-IRAK4 antibody [Y279] (ab32511) at 1/1000 dilution

Lanes 1 - 3:

Western blot - Anti-IRAK4 antibody [Y279] - BSA and Azide free (<a href='/en-us/products/primary-antibodies/irak4-antibody-y279-bsa-and-azide-free-ab239819'>ab239819</a>) at 1/1000 dilution

Lane 1:

MCF7 Input at 10 µg

Lane 2:

MCF7 Pulldown with ab32511 at 10 µL

Lane 3:

MCF7 Pulldown with Isotype Control <a href='/en-us/products/primary-antibodies/rabbit-igg-monoclonal-epr25a-isotype-control-ab172730'>ab172730</a> at 10 µL

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: 52 kDa

Observed band size: 55 kDa

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Exposure time: 45s