Intracellular IL-21 Staining Protocol

For MHC class-I-restricted GP_33–41 peptide (2 µg/ml) or MHC class-II restricted GP_67–77 (5 µg/ml) stimulation and staining were carried out as we have previously described (31 (link)). For polyclonal stimulation we used PMA (10 ng/ml) and ionomicyn (0.5 µg/ml) in place of peptide. For intracellular IL-21 staining, cells were permeabilized with saponin and incubated with 1:25 dilution of mouse IL-21R-human Fc (R&D Systems) for 30 minutes at 4°C, washed twice and stained with 1:200 anti-human Fc-PE (BD Pharmingen).

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Harker J.A., Wong K.A., Dolgoter A, & Zuniga E.I. (2015). Cell-intrinsic gp130 Signaling on CD4+ T cells Shapes Long-lasting Antiviral Immunity. Journal of immunology (Baltimore, Md. : 1950), 195(3), 1071-1081.

Publication 2015

mouse IL-21R-human Fc anti-human Fc-PE

Cells Dilution Fc pe Gp67 Human Il 21 Il 21r Intracellular Mhc class ii Mhc i peptide Mouse Peptide Saponin

Corresponding Organization :

Other organizations : University of California, San Diego

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Variable analysis

independent variables

MHC class-I-restricted GP33–41 peptide (2 µg/ml)
MHC class-II restricted GP67–77 (5 µg/ml)
PMA (10 ng/ml) and ionomycin (0.5 µg/ml)

dependent variables

Intracellular IL-21 staining

control variables

Staining carried out as previously described (31)

positive controls

PMA (10 ng/ml) and ionomycin (0.5 µg/ml)

negative controls

Not explicitly mentioned

Annotations

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