Citation Details

IFN regulatory factor-1 negatively regulates CD4+ CD25+ regulatory T cell differentiation by repressing Foxp3 expression

Authors	Fragale A, Gabriele L, Stellacci E, Borghi P, Perrotti E, Ilari R, Lanciotti A, Remoli AL, Venditti M, Belardelli F, Battistini A
In	J Immunol (2008) 181(3): 1673-82
Genes of Interest	Foxp3
Cells used in publication	T cell, unstim., human    Human unstimulated T cells    Blood/Immune Cells    Primary Cells    Species: human    Tissue Origin: blood T cell, mouse - C57BL/6    Mouse (C57BL/6) unstimulated T cells    Blood/Immune Cells    Primary Cells    Species: mouse    Tissue Origin: blood
Substrate	Plasmid (general)
Application	Co-Transfection
Topics	Luciferase Promoter analysis Signal transduction
Research Area	Immunology / Hematology
Relevant Products	Nucleofector Device Human T Cell Nucleofector Kit Mouse T Cell Nucleofector Kit
Reporter Genes	eGFP Renilla Luciferase
Vector Backbone	pEGFP

Nucleofection Experiments

Primary CD4 T cells in PBS supplemented with 0.5% BSA were incubated overnight at 4°C then 2.5 µg of the Foxp3 promoter luciferase reporter vector, 2 µg of IRF-1 expression vector, 0.2 µg of Renilla luciferase control vector, and 0.5 µg of pEGFP plasmid were added to 5x10^6 CD4 T cells resuspended in 100 µl of Nucleofector solution (Amaxa) and electroporated using the U-014 and X-001 program of the Nucleofector specific for untouched human and murine CD4 T cells, respectively. Transfection efficiency was monitored in all samples by FACS analysis of GFP fluorescence and was ranging from 30 to 50% for human cells depending on the donor in three independent experiments and from 15 to 25% for mouse cells. After 24 h, luciferase activity was measured.

Cell/Experiment Datasets

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