Detecting Antibody for ELISA to Analyze Mouse TNFa

October 19, 2015

Microbiology and Immunology
Loyola University Chicago
Research Associate

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Company:

BioLegend

Product Name:

Biotin anti-mouse TNF-α Antibody

Catalog Number:

506312

Info: View Product View Product Specs

Browse Similar Products: TNF-alpha Antibodies

We have isolated fractions from commensal bacteria secretions. To test if the secretions are able to suppress inflammatory activation, we measured the secretion of TNFa in mouse macrophage cells after LPS stimulation in the presence or absence of the fractions. We quantified the concentration of the TNFa using an antibody pair and standard from BioLegend. These reagents work pretty well with high sensitivity and reasonable price.

Experimental Design and Results Summary

Applications

ELISA

Sample

Cell culture supernatant

Primary Incubation

Capture antibody was coated (1.5 ug/ml ) overnight and incubate with standard protein for 1 hr at 37 degrees

Blocking Agent

5% nonfat milk in PBST

Secondary Incubation

0.5ug/ml of detecting antibody was added in each well and incubated for 1hr at 37 degrees

Tertiary Incubation

HPR-streptavidin was added and incubated for 1 hr at 37 degrees (1 : 10,000)

Detection

TMB was used to develop the signal

Results Summary

Using the antibody pair, the ELISA generated good signal that is sensitive enough to detect TNFa standard protein as low as 1 pg/ml. Using this we found that a fraction from the bacterial secretion suppressed LPS induced TFNa secretion in macrophages. We were able to detect around 20ug/ml in some of our samples when using to detect antibody at 0.5ug/ml concentration, so it is sensitive.

Additional Notes

This product has reasonable price, and easy to use

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Summary

The Good

Good price and can be used at lower concentration

The Bad

None

The Bottom Line

This is a very good antibody that I recommend to any lab who wants to use it to test mouse TNFa by ELISA. 0.5ug/ml can generate high sensitivity. I haven't test it in any other purpose except for ELISA.

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