LC3A/B human antibody, one of the best autophagy markers!

January 08, 2015

Biological Sciences
Rutgers University
Assist. Professor

Overall

Quality of Results

Ease-of-Optimization

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Company:

Cell Signaling Technologies

Product Name:

LC3A/B

Catalog Number:

12741

Browse Similar Products: LC3A/B Antibodies

The CST LC3A/B antibody is one of the most widely used autophagy markers on the market. It is usable in immunoblot assays in which the detection of the slightly lower band (also called LC3II) indicates the lipidation and autophagy induction (phagophore formation). Indeed, it is the antibody to use for the autophagy flux assay in which autophagy function is ascertained by induction of the autophagy pathway and lysosomal inhibition. In general, the best results are achieved using the ECL-based immunoblot assays and antibody concentrations of 1:1000. The antibody is also reliably working in immunofluorescence analyses at a concentration of 1:300 - here, dotted staining indicates LC3 lipidation and autophagosome buildup.

Experimental Design and Results Summary

Applications

Western Blot

Sample

MEFs, HeLa, 3T3, 293T, hiPSCs, primary neurons

Primary Incubation

1:1000, 4C, over night

Blocking Agent

5% BSA

Secondary Incubation

1:3000, RT, 2h

Tertiary Incubation

5% milk powder

Detection

ECL works best

Results Summary

Strong signals appear after an exposure of 30 sec to 1 min, depending on the ECL detection kit. The primary antibody can be reused multiple times.

Additional Notes

The blot shows the results from 293T cells in control conditions or treated with the lysosomal inhibitor Chloroquine or the mTOR inhibitor Torin1.

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Summary

The Good

It works in many tissues and cell lines.

The Bad

Has to use ECL for best results, does not work well on LiCoR

The Bottom Line

A great antibody for biochemistry and microscopy - highly recommended for everyone new to the autophagy field and needs to check for metabolic state of the analysed cells. See description for more information about how to read the autophagy flux assays and immunofluorescence signals. Good luck!

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