Identifies alpha Tubulin in Western Blots

October 06, 2015

Biological Sciences
Grambling State University
Assistant Professor

Overall

Quality of Results

Ease-of-Optimization

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Company:

Santa Cruz Biotechnology

Product Name:

α tubulin antibody

Catalog Number:

sc-5286

We carry out subcellular fractionation and verify the isolated fractions by western blot analysis. We use α tubulin as a cytosolic marker.

Experimental Design and Results Summary

Applications

Western Blot

Sample

Rat liver tissue and hepatoma cell lines

Primary Incubation

1:1000 overnight at 4C in 1:2 Odyssey buffer:TBS, 0.1% TWEEN 20

Blocking Agent

1:2 Odyssey buffer:TBS

Secondary Incubation

1:2 Odyssey buffer:TBS, 0.1% TWEEN 20, 0.02% SDS

Tertiary Incubation

NA

Detection

LiCor Odyssey

Results Summary

We see enrichment of α tubulin in the cytosolic fraction compared to the total lysate. The band migrates slightly below the 55 KDa molecular weight.

Additional Notes

We detect a nonspecific band at 17 KDa; something to consider for multiplexing.

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Summary

The Good

Works well at a reasonable 1:1000 dilution

The Bad

Some performance inconsistency

The Bottom Line

It has worked well for the most part. α tubulin should be easy. We experienced poor signal strength with a particular vial (well within the one year stability period) that cost us time and effort. The issue was resolved with a new vial but the rating reflects this inconsistency.

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