Detection of TNFa Secretion in Cells

Microbiology and Immunology
Loyola University Chicago
Research Associate

Overall

Quality of Results

Ease-of-Optimization

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Company:

BioLegend

Product Name:

Biotin anti-human TNFa

Catalog Number:

502904

To analyze macrophage activation, THP1 cells were differentiated into macrophage cells and treated with fraction 10 or control overnight. Cells were stimulated with LPS for 4 hours and then cell culture supernatant was collected for detecting TNFa secretion by ELISA. We found fraction treatment inhibited LPS induced TNFa secretion in macrophages.

Experimental Design and Results Summary

Applications

ELISA

Sample

Macrophage cell culture supernatant

Primary Incubation

Serially diluted samples or standard protein were added in a capture antibody coated microplate.

Blocking Agent

5% nonfat milk in PBST

Secondary Incubation

Biotin anti-human TNFa antibody (1ug/ml) was added in plate and incubated at 37 degrees for 1 hr

Tertiary Incubation

HPR-streptavidin

Detection

TMB

Results Summary

We found that the fraction from bacteria secretion-suppressed LPS induced TNFa secretion in human macrophages. The capture antibody is sensitive and easy to use. We usually combine capture antibody and protein standard from BioLegend for quantifying TNFa in our experiments. It is sensitive enough to detect TNFa standard protein as low as 1 pg/ml.

Additional Notes

None

Related Categories

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Summary

The Good

Using the capture and detection antibody pair from Biolegend, the ELISA generated good signal.

The Bad

None

The Bottom Line

The antibody is sensitive and specific for detection/quantification of human TNFa in cell culture supernatant.

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