Anti-IRAK-1 antibody [EPR19667] - BSA and Azide free

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-IRAK-1 antibody [EPR19667] - BSA and Azide free (AB251482)

This data was developed using ab208009, the same antibody clone in a different buffer formulation.

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cell line from cervix adenocarcinoma) cells labeling IRAK-1 with ab208009 at 1/100 dilution, followed by Goat Anti-Rabbit IgG (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing mainly cytoplasmic staining on HeLa cell line. The nuclear counter stain is DAPI (blue).

Tubulin is detected with Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor^® 594) (ab150120) secondary antibody at 1/1000 dilution (red).

The negative controls are as follows :

-ve control 1 : ab208009 at 1/100 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor^® 594) (ab150120) secondary at 1/1000 dilution.

-ve control 2 : Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary at 1/1000 dilution.

• WB

Supplier Data

Western blot - Anti-IRAK-1 antibody [EPR19667] - BSA and Azide free (AB251482)

This data was developed using ab208009, the same antibody clone in a different buffer formulation.

Blocking and dilution buffer : 5% NFDM/TBST.

Exposure time : Lanes 1, 3, and 4 : 1 minute; Lane 2 : 30 seconds.

There are two splice variants for IRAK-1, IRAK-1b and IRAK-1c,respectively. The product recognizes full length IRAK-1 (80 kDa) and IRAK-1c (68 kDa) (PMID : 16690127).

All lanes:

Western blot - Anti-IRAK-1 antibody [EPR19667] (<a href='/en-us/products/primary-antibodies/irak-1-antibody-epr19667-ab208009'>ab208009</a>) at 1/1000 dilution

Lane 1:

HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 10 µg

Lane 2:

HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate at 10 µg

Lane 3:

HepG2 (Human liver hepatocellular carcinoma cell line) whole cell lysate at 10 µg

Lane 4:

A549 (Human lung carcinoma 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: 76 kDa

Observed band size: 68 kDa,80 kDa

false

• WB

Lab

Western blot - Anti-IRAK-1 antibody [EPR19667] - BSA and Azide free (AB251482)

This data was developed using ab208009, the same antibody clone in a different buffer formulation.

Lane 1 : Wild type HAP1 whole cell lysate (40 μg)

Lane 2 : IRAK-1 knockout HAP1 whole cell lysate (40 μg)

Lane 3 : HeLa whole cell lysate (40 μg)

Lane 4 : HEK293 whole cell lysate (40 μg)

Lanes 1 - 4 : Merged signal (red and green). Green - ab208009 observed at 85 kDa. Red - loading control, ab8245, observed at 37 kDa.

ab208009 was shown to specifically react with IRAK-1 when IRAK-1 knockout samples were used. Wild-type and IRAK-1 knockout samples were subjected to SDS-PAGE. ab208009 and ab8245 (Mouse anti GAPDH loading control) were incubated overnight at 4°C at 1/200 dilution and 1/10000 dilution respectively. Blots were developed with 800CW Goat anti Rabbit and 680CW Goat anti Mouse secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-IRAK-1 antibody [EPR19667] (<a href='/en-us/products/primary-antibodies/irak-1-antibody-epr19667-ab208009'>ab208009</a>)

Predicted band size: 76 kDa

false