GNAS Antibody for Western Blotting

January 28, 2016

Biochemistry
Stanford University
Senior Research Scientist

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Company:

Everest Biotech

Product Name:

Goat Anti-GNAS Antibody

Catalog Number:

EB10063

Browse Similar Products: GNAS Western blot Antibodies

The Gα s subunit of heterotrimeric G proteins stimulates adenylate cyclases to generate cAMP, an important second messenger molecule that is necessary for PKA activation. The goal of this experiment is to test if NIH3T3 cells express Gα s (a protein product of the GNAS gene). If it is expressed, I would then go on to determine the localization of this protein. This is the first antibody that I have purchased to test its performance in Western Blotting due to the clean image supplied by the manufacturer.

Experimental Design and Results Summary

Applications

Western Blot

Sample

NIH3T3 whole cell extract

Primary Incubation

1:1600 in blocking agent for 16 hr at 4 deg

Blocking Agent

5% Milk powder in 1x TBS-T (0.1% Tween-20)

Secondary Incubation

Donkey anti-goat IgG HRP at 1:5000 dilution in blocking agent for 1 hr at room temp

Tertiary Incubation

None

Detection

ECL pico for 5 min

Results Summary

This antibody detects a nice band at 46 kDa (predicted size) with no background bands.

Additional Notes

None

Related Categories

Image Gallery

Summary

The Good

Specific band detection

The Bad

Signal can be weak sometimes

The Bottom Line

It worked very well for the first two times but a decline in performance was observed later. It could be due to repeated free thaw cycles. I would recommend following manufacturer recommendation of avoiding free-thaw cycles even if it was for few rounds. One should prepare small aliquots for one time use.

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