Myc tag antibody for Western Blotting

February 04, 2016

Biochemistry
Stanford University
Senior Research Scientist

Overall

Quality of Results

Ease-of-Optimization

What do these ratings mean?
Write a Review

Company:

Sigma-Aldrich

Product Name:

Anti-c-myc

Catalog Number:

11667149001

I am interested in understanding the mechanism of Hedgehog signaling. My project involves protein biochemistry techniques like protein purification, Western Blotting, enzymatic assays etc. The goal of this experiment is to test if Myc tag antibody from Sigma can be used for Western Blotting experiments of a Myc-tagged protein over-expressed in HEK293T cells. This antibody was selected based on a recommendation from a fellow researcher.

Experimental Design and Results Summary

Applications

Western Blot

Sample

HEK293T whole cell extract

Primary Incubation

1:2000 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-mouse IgG HRP at 1:5000 dilution in blocking agent for 1 hr at room temp.

Tertiary Incubation

None

Detection

ECL for 1 min on a film

Results Summary

A 3xMyc-tagged EFCAB7 construct was over-expressed in HEK293T cells for 24 hours. Whole cell extract was prepared in RIPA lysis buffer and 50 µg of protein was loaded for SDS-PAGE followed by Western blotting with Myc antibody from Sigma. Myc-EFCAB7 should run at ~80 kDa and this antibody nicely picks up the signal.

Additional Notes

None

Related Categories

Image Gallery

Summary

The Good

Good signal for a 3x Myc tag

The Bad

Low signal might be obtained sometimes with a 1x Myc tag (especially if it is an internal tag)

The Bottom Line

Good antibody for detection of either N or C terminal Myc tagged proteins in Western Blotting

Share your experience with other scientists. Write a Review! »

Join the discussion