Anti-MyD88 antibody [EPR21824]

• IP

Supplier Data

Immunoprecipitation - Anti-MyD88 antibody [EPR21824] (AB219413)

MyD88 was immunoprecipitated from 0.35 mg RAW 264.7 (Mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysate with ab219413 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab219413 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/5000 dilution.

Lane 1 : RAW 264.7 whole cell lysate 10 μg (Input).
Lane 2 : ab219413 IP in RAW 264.7 whole cell lysate (+).
Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab219413 in RAW 264.7 whole cell lysate (-).
Blocking/Dilution buffer : 5% NFDM/TBST.
Exposure time : 3 minutes.

All lanes:

Immunoprecipitation - Anti-MyD88 antibody [EPR21824] (ab219413)

Predicted band size: 33 kDa

false

• WB

Lab

Western blot - Anti-MyD88 antibody [EPR21824] (AB219413)

Blocking/Dilution buffer : 5% NFDM/TBST.

ab219413 was shown to specifically react with MyD88 in wild-type HAP1 cells as signal was lost in MyD88 knockout cells. Wild-type and MyD88 knockout samples were subjected to SDS-PAGE. ab219413 and ab181602 (Rabbit anti-GAPDH loading control) were incubated 1 hour at room temperature at 1/1000 dilution and 1/200,000 dilution respectively. Blots were developed with Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) secondary antibody at 1/100,000 dilution for 1 hour at room temperature before imaging. The blot was developed on a BIO-RAD^® ChemiDoc™ MP instrument using the ECL technique

All lanes:

Western blot - Anti-MyD88 antibody [EPR21824] (ab219413) at 1/100000 dilution

Lane 1:

Wild-type HAP1 whole cell lysate at 20 µg

Lane 2:

MyD88 knockout HAP1 whole cell lysate at 20 µg

Lane 3:

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

Lane 4:

Ramos (Human Burkitt's lymphoma B lymphocyte) 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: 33 kDa

false

Exposure time: 3min

• WB

Supplier Data

Western blot - Anti-MyD88 antibody [EPR21824] (AB219413)

False colour image of Western blot : Anti-MyD88 antibody [EPR21824] staining at 1/1000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab219413 was shown to bind specifically to MyD88. A band was observed at 35 kDa in wild-type A549 cell lysates with no signal observed at this size in MYD88 knockout cell line ab286715 (knockout cell lysate ab290793). To generate this image, wild-type and MYD88 knockout A549 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % 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 IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

All lanes:

Western blot - Anti-MyD88 antibody [EPR21824] (ab219413) at 1/1000 dilution

Lane 1:

Wild-type A549 cell lysate at 20 µg

Lane 2:

MYD88 knockout A549 cell lysate at 20 µg

Lane 2:

Western blot - Human MYD88 knockout A549 cell line (<a href='/en-us/products/cell-lines/human-myd88-knockout-a549-cell-line-ab286715'>ab286715</a>)

Lane 3:

HEK-293 cell lysate at 20 µg

Secondary

Lanes 1 - 3:

Goat anti-Mouse IgG H&L 680RD

Lanes 1 - 3:

Goat anti-Rabbit IgG H&L 800CW

Predicted band size: 33 kDa

Observed band size: 35 kDa

false

• WB

Supplier Data

Western blot - Anti-MyD88 antibody [EPR21824] (AB219413)

Blocking/Dilution buffer : 5% NFDM/TBST

All lanes:

Western blot - Anti-MyD88 antibody [EPR21824] (ab219413) at 1/1000 dilution

Lane 1:

Rat liver lysate at 20 µg

Lane 2:

RAW 264.7 (Mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysate at 20 µg

Lane 3:

A20 (Mouse reticulum sarcoma B lymphocyte) cell lysate at 20 µg

Lane 4:

Mouse liver lysate at 20 µg

Lane 5:

Mouse lung 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: 33 kDa

false

Exposure time: 3min