The PBMCs were isolated by density gradient centrifugation, using the Ficoll-Hypaque technique ( Solarbio, China), and freshly used for surface and intracellular staining to analyze frequency, phenotype, and function. To detect cytokine production and cytotoxicity markers, PBMCs were cultured in RPMI-1640 medium supplemented with 10% fetal bovine serum, and stimulated with phorbol 12-myristate-13-acetate(PMA)/ionomycin and Brefeldin A/Monensin (Multi sciences, China) for 4–6 h at 37 °C. Stimulated PBMCs were fixed and permeabilized using a FIX & PERM Kit (Multi sciences, China) to stain the intracellular markers. The PBMCs were further stimulated with IL-8, IL-12, or IL-18 (Novoprotein Scientific, China) at 50 ng/mL for 24 h.
Pma ionomycin and brefeldin a monensin
Manufactured by MultiSciences Biotech
Sourced in China
PMA)/ionomycin is a combination of the phorbol ester phorbol 12-myristate 13-acetate (PMA) and the calcium ionophore ionomycin. This combination is commonly used to stimulate and activate T cells and other immune cells in vitro. Brefeldin A is a lactone that interferes with protein transport from the endoplasmic reticulum to the Golgi apparatus, leading to the accumulation of proteins within the endoplasmic reticulum. Monensin is a carboxylic ionophore that disrupts the function of the Golgi apparatus, also leading to the accumulation of proteins. These two compounds, Brefeldin A and Monensin, are often used together to study protein secretion and trafficking in cells.
Automatically generated - may contain errors
Lab products found in correlation
2 protocols using pma ionomycin and brefeldin a monensin
1
Immunophenotyping and Cytokine Analysis of PBMCs
2
PBMC Isolation and Functional Assays
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The PBMCs were isolated by density gradient centrifugation, using the Ficoll-Hypaque technique ( Solarbio, China), and freshly used for surface and intracellular staining to analyze frequency, phenotype, and function. To detect cytokine production and cytotoxicity markers, PBMCs were cultured in RPMI-1640 medium supplemented with 10% fetal bovine serum, and stimulated with phorbol 12-myristate-13-acetate(PMA) /ionomycin and Brefeldin A/Monensin (Multi sciences, China) for 4 -6 h at 37 °C. Stimulated PBMCs were xed and permeabilized using a FIX & PERM Kit (Multi sciences, China) to stain the intracellular markers. The PBMCs were further stimulated with IL-8, IL-12, or IL-18 (Novoprotein Scienti c, China) at 50ng/mL for 24 h.
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