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Sample GSM845889 Query DataSets for GSM845889
Status Public on Apr 10, 2013
Title P1
Sample type RNA
 
Source name CD4+T cells stimulated by PHA/IL-2 for 6 days
Organism Homo sapiens
Characteristics cell type: human CD4+T cells
Treatment protocol Fresh normal donors peripheral blood mononuclear cells isolated by Ficool-Hypaque (TBD sciences) centrifugation were enriched for CD4+ T cells (>97% purity by FACS) by magnetic negatively selection (CD4+T cell Isolation Kit II, Miltenyi Biotec). For PHA/IL-2 stimulation, cells were cultured at a density of 2×106/well with human recombinant IL-2 (100 units/ml, Pepro Tech) and PHA (5µg/ml, Sigma). For CD3/CD28 costimulation, cells were cultured at a density of 0.5×106/well with polystyrene beads coated of CD3 and CD28 antibodies (Dynabeads CD3/CD28 T Cell Expander, Dynal). The ratio of beads to cells was 3:1.
Growth protocol Cells were cultured in 6-well plates at 37℃, 5% CO2 in RPMI 1640 medium supplemented with 10% heat-inactivated fetal bovine serum and 20mM HEPES
Extracted molecule total RNA
Extraction protocol Total RNA was extracted using TRIZOL Reagent (Cat#15596-018,Life technologies, Carlsbad, CA, US)following the manufacturer’s instructions and checked for a RIN number to inspect RNA integration by an Agilent Bioanalyzer 2100 (Agilent technologies, Santa Clara, CA, US).Qualified total RNA was further purified by RNeasy mini kit (Cat#74106, QIAGEN, GmBH, Germany) and RNase-Free DNase Set (Cat#79254, QIAGEN, GmBH, Germany).
Label Cy3
Label protocol Total RNA was amplified and labeled by Low Input Quick Amp Labeling Kit, One-Color (Cat#5190-2305, Agilent technologies, Santa Clara, CA, US), followed the manufacturer’s instructions. Labeled cRNA were purified by RNeasy mini kit (Cat#74106, QIAGEN, GmBH, Germany).
 
Hybridization protocol Each Slide was hybridized with 1.65μg Cy3-labeled cRNA using Gene Expression Hybridization Kit (Cat#5188-5242, Agilent technologies, Santa Clara, CA, US) in Hybridization Oven (Cat#G2545A, Agilent technologies, Santa Clara, CA, US), according to the manufacturer’s instructions. After 17 hours hybridization, slides were washed in staining dishes (Cat#121, Thermo Shandon, Waltham, MA, US) with Gene Expression Wash Buffer Kit(Cat#5188-5327, Agilent technologies, Santa Clara, CA, US), followed the manufacturer’s instructions.
Scan protocol Slides were scanned by Agilent Microarray Scanner (Cat#G2565CA, Agilent technologies, Santa Clara, CA, US) with default settings,Dye channel: Green ,Scan resolution=5μm, PMT 100%,10% ,16bit.
Description Gene expression of CD4+T cells stimulated by PHA/IL-2 for 6 days
Data processing The scanned images were analyzed with Feature Extraction software (Agilent technologies, Santa Clara, CA, US)with default settings.
 
Submission date Dec 08, 2011
Last update date Apr 10, 2013
Contact name Hua-Tang Zhang
E-mail(s) zhanght@mail.kiz.ac.cn
Organization name Kunming Institute of Zoology, Chinese Academy of Science
Department key laboratory of animal models and human diseases mechanisms
Lab the Laboratory of Immunobiology
Street address No.32 Jiaochang Donglu
City Kunming
State/province Yunnan
ZIP/Postal code 650223
Country China
 
Platform ID GPL4133
Series (1)
GSE34252 CD4+T cells activation

Data table header descriptions
ID_REF
VALUE processed Cy3 signal intensity (Agilent gProcessedSignal)

Data table
ID_REF VALUE
1 2.280278e+004
2 2.793666e+000
3 2.796098e+000
4 4.150564e+000
5 2.801855e+000
6 2.805260e+000
7 2.808954e+000
8 2.812831e+000
9 2.817216e+000
10 2.821738e+000
11 2.826553e+000
12 8.321084e+002
13 4.406429e+002
14 2.808858e+003
15 1.551209e+002
16 3.541751e+004
17 8.899381e+001
18 2.253475e+002
19 1.261421e+005
20 1.132662e+002

Total number of rows: 45015

Table truncated, full table size 868 Kbytes.




Supplementary file Size Download File type/resource
GSM845889_251485049075_S01_GE1_105_Dec08_1_2.txt.gz 9.1 Mb (ftp)(http) TXT
Processed data included within Sample table
Processed data provided as supplementary file

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