Cytokine Analysis and NK Cell Proliferation

Cytokine production and degranulation – as measured by the appearance of LAMP1/CD107a at the cell surface and accumulation of intracellular cytokines, was performed on freshly isolated PBMC and analyzed on a flow cytometer (LSR II, BD Biosciences), gating on live CD56+ CD3- cells to identify NK cells, as described (dx.doi.org/10.17504/protocols.io.8epv51me6l1b/v1). Proliferation of NK cells in enriched populations or amongst PBMC were measured by the dilution of a permanent membrane dye by flow cytometry (dx.doi.org/10.17504/protocols.io.261geojwol47/v1).

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Kennedy P.R., Vallera D.A., Ettestad B., Hallstrom C., Kodal B., Todhunter D.A., Bendzick L., Hinderlie P., Walker J.T., Pulkrabek B., Pastan I., Kratzke R.A., Fujioka N., Miller J.S, & Felices M. (2023). A tri-specific killer engager against mesothelin targets NK cells towards lung cancer. Frontiers in Immunology, 14, 1060905.

Publication 2023

Cells Cells proliferation Cytokines Dye dilution Flow cytometry Intracellular Lamp1 Membrane Nk cells Populations

Corresponding Organization : University of Minnesota

Other organizations : National Institutes of Health, National Cancer Institute

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Variable analysis

independent variables

Enrichment of NK cells in the populations

dependent variables

Cytokine production
Degranulation (as measured by the appearance of LAMP1/CD107a at the cell surface)
Accumulation of intracellular cytokines
Proliferation of NK cells (as measured by the dilution of a permanent membrane dye)

control variables

Freshly isolated PBMC
Gating on live CD56+ CD3- cells to identify NK cells

controls

Positive controls: Not explicitly mentioned
Negative controls: Not explicitly mentioned

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