Anti-MDA5 antibody [EPR6743] - BSA and Azide free

• WB

Unknown

Western blot - Anti-MDA5 antibody [EPR6743] - BSA and Azide free (AB219952)

This data was developed using ab126630, the same antibody clone in a different buffer formulation.

All lanes:

Western blot - Anti-MDA5 antibody [EPR6743] (<a href='/en-us/products/primary-antibodies/mda5-antibody-epr6743-ab126630'>ab126630</a>) at 1/1000 dilution

All lanes:

THP1 treated with LPS cell lysate at 10 µg

Secondary

All lanes:

Goat anti-rabbit HRP conjugated antibody at 1/2000 dilution

Predicted band size: 117 kDa

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• WB

Lab

Western blot - Anti-MDA5 antibody [EPR6743] - BSA and Azide free (AB219952)

Western blot : Anti-IFIH1 antibody [EPR6743] (ab126630) 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, ab126630 was shown to bind specifically to IFIH1. A band was observed at 140, 90 kDa in wild-type Jurkat cell lysates with no signal observed at this size in IFIH1 knockout cell line. To generate this image, wild-type and IFIH1 knockout Jurkat 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. This data was developed using ab126630, the same antibody clone in a different buffer formulation.

All lanes:

Western blot - Anti-MDA5 antibody [EPR6743] (<a href='/en-us/products/primary-antibodies/mda5-antibody-epr6743-ab126630'>ab126630</a>) at 1/1000 dilution

Lane 1:

Wild-type Jurkat, LPS (1 ug/mL, 24 h) cell lysate at 20 µg

Lane 2:

Wild-type Jurkat, vehicle control LPS (0 ug/mL, 24 h) cell lysate at 20 µg

Lanes 3 - 4:

Western blot - Human IFIH1 knockout Jurkat cell line (<a href='/en-us/products/cell-lines/human-ifih1-knockout-jurkat-cell-line-ab305280'>ab305280</a>)

Lane 3:

IFIH1 knockout Jurkat, LPS (1 ug/mL, 24 h) cell lysate at 20 µg

Lane 4:

IFIH1 knockout Jurkat, vehicle control LPS (0 ug/mL, 24 h) cell lysate at 20 µg

Predicted band size: 117 kDa,90 kDa

Observed band size: 140 kDa

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• WB

Lab

Western blot - Anti-MDA5 antibody [EPR6743] - BSA and Azide free (AB219952)

This data was developed using the same antibody clone in a different buffer formulation (ab126630)

ab126630 was shown to react with IFIH1 in wild-type DMS 53 cells in Western blot with loss of signal observed in a IFIH1 siRNA knockdown cell line. Cell lysates from wild-type DMS 53 transfected with either scrambled siRNA or IFIH1 siRNA were subjected to SDS-PAGE. Membranes were blocked in 5% milk in TBST for 1 hr before incubation with ab126630 overnight at 4 °C at a 1/1000 dilution. Blots were incubated with secondary antibodies at 0.2 µg/mL before imaging.

This data was provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.

All lanes:

Western blot - Anti-MDA5 antibody [EPR6743] (<a href='/en-us/products/primary-antibodies/mda5-antibody-epr6743-ab126630'>ab126630</a>) at 1/1000 dilution

Lane 1:

Wild-type DMS 53 transfected with scrambled siRNA control lysate at 30 µg

Lane 2:

DMS 53 transfected with siRNA specifically targeting IFIH1 cell lysate at 30 µg

Observed band size: 116 kDa

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