Mouse mAb Against Human Neutrophil Elastase

The Cell and Molecular Biology
The Scripps Research Institute
Assistant Professor

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Company:

Santa Cruz Biotechnology

Product Name:

Neutrophil Elastase Antibody

Catalog Number:

sc-55549

The detection of human neutrophil elastase (NE) is important in enzymology, where NE release from neutrophils or its presence should be confirmed and quantified.The antibody sc-55549 was designed to detect human neutrophil elastase in western blotting and immunofluorescence. We used this antibody to compare in the same samples the total amount of NE released from human neutrophils versus NE activity (determined by substrate cleavage assay).

Experimental Design and Results Summary

Applications

Western Blot

Sample

Purified neutrophil elastase, 1 microgram; U937 cell lsate, 20 micrograms; human neutrophils, 2x10e^5 cells

Primary Incubation

1 ug/ml in PBS supplemented with 5% nonfat milk and 0.05% Tween 20; overnight, cold room

Blocking Agent

5% nonfat milk in PBS/0.05% Tween 20

Secondary Incubation

goat anti-mouse IgG conjugated with HRP; 1:1,000; 1 hr, room temperature

Tertiary Incubation

None

Detection

West pico

Results Summary

In our hands, this antibody did not recognize neutrophil elastase from any tested sources, including neutrophil elastase purified from human neutrophils loaded up to 1 microgram per lane, contents of 2x10e^5 human neutrophils, and cell lysate of U937 cells, which according to the company should provide positive control. The antibody failed to recognize neutrophil elastase (~29.5 kDa) under both reducing and non-reducing conditions. The image shows negative results under non-reducing conditions.

DOI or PMID #

N/A

Additional Notes

N/A

Related Categories

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Summary

The Good

None

The Bad

The antibody failed to reliably detect human neutrophil elastase in WB.

The Bottom Line

I would not recommend this antibody for detection of human neutrophil elastase by western blotting

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