Activation Cocktail for Cytokine Quantification by Flow Cytometry

August 15, 2023

BlueskyReddit
The Salk Institute of Biological Studies
NOMIS CENTER FOR IMMUNOBIOLOGY AND MICROBIAL PATHOGENESIS
Postdoctoral Researcher

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Quality of Results

Ease-of-Optimization

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Company:

BioLegend

Product Name:

Cell Activation Cocktail (with Brefeldin A)

Catalog Number:

423304

My research area includes immune phenotyping of primary or metastatic tumors. I often do ex vivo stimulation of T cells sorted from the tumor microenvironment. This activation cocktail is very helpful as it does not require additional steps of monensin or brefeldin A addition.

Experimental Design and Results Summary

Application

Activation of immune cells for cytokine quantification.

Starting Material

Cells from tissue or primary lymphoid organs.

Tips

4-4.5 hrs of activation is sufficient. It reduces the overall dead cell population.

Results Summary

The result here is shown for IFNg and TNF-a production ex vivo isolated T cells from the metastatic tumor and activated for 4.5 hrs using BioLegend activation cocktail mix. The cocktail mix was used at 1:500 dilution for 0.1 million immune cells activated in 0.5 ml complete RPMI. After 4.5 hours of activation, cells were harvested and stained with a fixable viability dye and various antibodies. The result shown here is for IFNg and TNFa production by live-gated CD45+CD4+TCRb+ T cells.

DOI or PMID #

N/A

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Summary

The Good

Does not require additional step of Brefeldin-A or Monensin and saves a lot of time.

The Bad

If kept for long time (over 4.5hrs), the dead cell population increases.

The Bottom Line

Easy to prepare the culture media with activating cocktails, less calculation, and saves more time. I would definitely use it.

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