Sepharose G Beads for Immunoprecipitation Application

November 02, 2015

BlueskyReddit
Texas Tech University Health Science Center
Internal Medicine
Postdoctoral Researcher

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Company:

GE Healthcare

Product Name:

Protein G Sepharose 4 Fast Flow

Catalog Number:

17-0618-01

These beads are used to enrich a protein of low abundance by the pull down method. Basically it binds according to the IgG, upon the source of the particular immunoglobulin.

Experimental Design and Results Summary

Application

Western blot/Immunoprecipitation

Starting Material

Mouse cell lysate

Protocol Overview

G beads are used to pull down a particular protein of interest from the whole cell lysate. The whole cell lysate was incubated overnight with the particular antibody and the next day, the beads were added to it according to the source of the antibody and incubated again for 2hrs. After incubation, it was centrifuged at lower RPM, washed two times with RIPA buffer and loaded for western blot.

Tips

This G Sepharose beads are strong for rabbit IgG and mouse IgG1 and weak for other IgG sources.

Results Summary

Protein G binds specifically to the Fc portion of IgG. Based on the source of the antibody, either the Heavy chain or light chain of the IgG will be detected.

Additional Notes

None

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Summary

The Good

It is a straightforward method and the results are very accurate.

The Bad

Sometimes it will give dark background and if your protein of interest is the same as the size of the IgG, then it is hard to predict.

The Bottom Line

Protein G beads are most commonly used in immunoprecipitation assays and used to enrich proteins of low abundance in your whole cell lysate.

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