Less-than-Ideal Anti-Tubulin Antibody

Physiology
University of Tennessee HSC
Postdoctoral Researcher

Overall

Quality of Results

Ease-of-Optimization

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Company:

Santa Cruz Biotechnology Inc

Product Name:

Tubulin (5F131)

Catalog Number:

sc-73242

Clone Number:

5F131

Human cells were growth-arrested, treated with a specific growth factor; protein extracts were made and resolved in a 10% SDS PAGE. Resolved proteins were western transferred in the nitrocellulose membrane and immunodetected for STAT1 protein levels. For normalisation of the blot, the same membrane was reprobed with anti- Tubulin antibody which was obtained from Santacruz Biotechnology Inc. Selection of antibody was based on the information provided in the flyer.

Experimental Design and Results Summary

Applications

Western Blot

Sample

Human cell protein extract

Primary Incubation

Dilution (1:500) in 5% skimmed milk prepared in 1x TBST, overnight incubation at 4 degree Celsius

Blocking Agent

5% skimmed milk made in 1x TBST

Secondary Incubation

Goat anti-mouse IgG HRP, dilution (1:4000) in 5% skimmed milk made in 1x TBST, 1hr at RT

Tertiary Incubation

None

Detection

Enhanced chemiluminescent detection in X-Ray films

Results Summary

Indicated anti-tubulin antibody from Santacruz Biotechnology Inc has detected only one specific band of tubulin at the correct size. However, the signal was too low, and the membrane was full of patches may be due to some impurities, which leads to nonspecific binding of primary Ab to the nitrocellulose membrane. This conclusion was made by performing the same experiment at least 4 times with the same antibody.

DOI or PMID #

N/A

Additional Notes

None

Related Categories

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Summary

The Good

Gives single band

The Bad

A lot of non-specificity in the form of patches.

The Bottom Line

Poor antibody

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