9 protocols
AccessionNameType
P-GSE46185-1
normalization data transformation protocol
Expression values were determined using Affymetrix GeneChip Command Console Software (AGCC) and Console Software (Expression Console). ID_REF = VALUE = MAS5.0 signal intensity
P-GSE46185-7
array scanning protocol
GeneChips were scanned using the GeneChip Scanner 3000.
P-GSE46185-6
hybridization protocol
cRNA was hybridized to GeneChip Mouse Genome 430 2.0 Arrays from Affymetrix according the manufacturers protocol.
P-GSE46185-5
labelling protocol
Total RNA was labeled using the GeneChip 3' IVT Express Kit
P-GSE46185-4
nucleic acid extraction protocol
Trizol extraction of total RNA was performed according to the manufacturer's instructions.
P-GSE46185-9
growth protocol
Splenic CD4 T cells were stimulated under Th2 culture conditions for 5 days in vitro. The Th2 cells were further cultured in vitro for another 2 days in the absence of any exogenous cytokines. The cultured CD4 T cells were then restimulated with immobilized anti-TCR mAbs with IL-2 and anti-IL-4 mAbs for 5 days. This cycle was repeated three times. (Th2-4th cells)
P-GSE46185-8
sample treatment protocol
Th2-4th cells were collected and treated with control siRNA (AM4635; Applied Biosystems) or Gata3 siRNA (s66482; Applied Biosystems) using the Mouse T cell Nucleofector Kit (Amaxa) according to the manufacturer's protocol. Mock- or Gata3 siRNA-transfected Th2-4th cells were harvested after 24hrs, and stimulated with or without immobilized anti-TCR mAb (H57-597; 3 mg/ml) for 4hrs. Then cells were collected and resuspendet in the Trizol solution (GibcoBRL).
P-GSE46185-3
growth protocol
Splenic CD4 T cells were prepared using a magnetic cell sorter (AutoMACS; Miltenyi Biotec) yielding a purity of >98%. Where indicated, cells from C57BL/6 mice were stimulated with immobilized anti-TCR mAb (H57−597; 3 mg/ml) and anti-CD28 mAb under Th2-culture conditions for 2days in vitro. Th2 conditions; 25 U/ml IL-2, 100 U/ml IL-4. After culturing for 3 more days, the cells were harvested.
P-GSE46185-2
sample treatment protocol
Th2 culture cells were collected and treated with control siRNA (AM4635; Applied Biosystems) or Gata3 siRNA (s66482; Applied Biosystems) using the Mouse T cell Nucleofector Kit (Amaxa) according to the manufacturer's protocol. Mock- or Gata3 siRNA-transfected Th2 cells were harvested after 24hrs, and stimulated with or without immobilized anti-TCR mAb (H57-597; 3 mg/ml) for 4hrs. Then cells were collected and resuspendet in the Trizol solution (GibcoBRL).