Total IRE1alpha

Food Science and Human Nutrition
Colorado State University
Graduate Student

Overall

Quality of Results

Ease-of-Optimization

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Company:

Abcam

Product Name:

Anti-IRE1 antibody

Catalog Number:

ab37073

IRE1alpha is a key protein that is phosphorylated and activated during ER stress, initiating the unfolded protein response. Determining the activation state of IRE1alpha due to a variety of stress conditions is helpful in assessing the severity of ER stress.

Experimental Design and Results Summary

Applications

Western Blot

Sample

Rat hepatoma H4IIE liver cells

Primary Incubation

1:500 in 1:3 odyssey block:1X TBST (0.1% Tween) shaking, overnight at 4 degrees C

Blocking Agent

1:3 odyssey block:1X TBS at room temperature for 1 hr.

Secondary Incubation

1:15000 Goat anti-rabbit IRDye in 1:3 odyssey block:1X TBST (0.1% Tween) and 0.2% SDS for one hour at room temperature in the dark

Tertiary Incubation

N/A

Detection

Licor Odyssey

Results Summary

Total IRE1alpha is pretty clear at 125kDa but there is a ton of non-specific binding and two prominent bands near 40kDa that show up so strong they mask the signal from IRE1. Have to adjust the brightness and contrast of the image to see IRE1alpha.

Additional Notes

None

Related Categories

Image Gallery

Summary

The Good

Antibody works.

The Bad

A lot of background.

The Bottom Line

This is the first IRE1 antibody that has worked in our cell line. Would recommend for others if no other IRE1 antibody from other companies have worked.

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