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Sample GSM4053177 Query DataSets for GSM4053177
Status Public on Sep 04, 2020
Title Donor 1, Timepoint 8, Replicate A
Sample type RNA
 
Source name CD4+
Organism Homo sapiens
Characteristics gender: female
Treatment protocol 3.5x105 cells/well of the freshly isolated CD4+ cells were seeded into 100 µl of medium within a 96-well plate by using a multichannel pipette. Cells were subsequently activated in separate wells by αCD2/αCD3/αCD28 MACSiBead™ particles (T cell activation/expansion kit, human, Miltenyi Biotec GmbH, Bergisch Gladbach, Germany). Sampling during a total time period of 24 h was conducted, collecting cells of each donor from three separate wells at intervals of 2 h. Cells were pelleted and lysed immediately by QIAzol® Lysis Reagent from miRNeasy Micro Kit (Qiagen, Hilden, Germany). Samples were frozen (-70 °C) for less than 2 months before RNA extraction.
Growth protocol Venous blood samples were obtained from two age and gender matched volunteers (female; Donor 1: Age 26 years; Donor 2: Age 23 years). Samples were collected using lithium heparin containing vacutainer collection tubes (S-Monovette®, Sarstedt AG& Co. KG, Numbrecht, Germany). PBMCs were isolated by Ficoll density gradient centrifugation. CD4+ T cells were isolated by negative selection (Human CD4+ T cell Isolation Kit, Miltenyi Biotech, Bergisch Gladbach, Germany). Cells were resolved and cultured in RPMI 1640 medium (Life Technologies GmbH, Darmstadt, Germany), supplemented with 10 % heat inactivated fetal bovine serum (Biochrom GmbH, Berlin, Germany), penicillin (100 U/mL) and streptomycin (100 μg/mL)
Extracted molecule total RNA
Extraction protocol Total RNA was extracted in a semi-automated procedure, using QIAcube robot systems and following the manufacturers` instructions for miRNeasy Micro Kit (Qiagen, Hilden, Germany). The RNA was quantified by Qubit® Fluorometer using Qubit® RNA HS Assay Kit (Thermo Fisher Scientific Inc., Waltham, Massachusetts, United States). RNA integrity was verified using Agilent 2100 Bioanalyzer instrument and RNA Pico Kit (Agilent Technologies, Santa Clara, California, United States). To determine mRNA expression profiles, 25 ng total RNA of each sample was utilized within mRNA One-Color Microarray-Based Gene Expression analysis.
Label Cy3
Label protocol The synthesis of labeled cRNA was performed using Low Input Quick Amp Labeling Kit (One-Color) (Agilent Technologies, Santa Clara, California, United States). The cRNA samples were purified by RNeasy Mini Kit (Qiagen, Hilden, Germany) and concentrations of the purified samples were determined by NanoDrop™ 2000c Spectrophotometer (Thermo Fisher Scientific Inc., Waltham, Massachusetts, United States) following the instructions of the labeling kit.
 
Hybridization protocol Hybridization was performed using the Gene Expression Hybridization Kit and Human SurePrint G3 Gene Expression Microarrays (V3, G4851C; Agilent Technologies, Santa Clara, California, United States) following the manufacturer’s protocol.
Scan protocol Arrays were scanned with a resolution of 3 µm. Raw data was extracted using Feature Extraction software (Agilent Technologies, Santa Clara, California, United States).
Description Gene expression after T cell activation (Timepoint 8)
Data processing Raw expression values were background corrected, quantile normalized and log2 transformed using the limma R-package (method = normexp, offset = 16).
 
Submission date Aug 29, 2019
Last update date Sep 05, 2020
Contact name Tim Kehl
E-mail(s) tkehl@bioinf.uni-sb.de
Phone +49-681-302-68613
Organization name Saarland University
Department Center for Bioinformatics
Street address Campus E2.1, P.O. box 151150
City Saarbrücken
ZIP/Postal code 66041
Country Germany
 
Platform ID GPL13607
Series (1)
GSE136625 Time-resolved RNA atlas of early human T cell activation (mRNA)

Data table header descriptions
ID_REF
VALUE quantile

Data table
ID_REF VALUE
1 16.07854671
2 4.718272094
3 4.718272094
4 5.772382129
5 4.718272094
6 5.967987461
7 8.098268399
8 11.95236134
9 5.253944252
10 4.892234801
11 5.136593221
12 4.955989546
13 5.87583167
14 4.593343146
15 11.42165072
16 4.409216092
17 6.159499905
18 9.779704728
19 5.324913547
20 7.243209656

Total number of rows: 62976

Table truncated, full table size 1090 Kbytes.




Supplementary file Size Download File type/resource
GSM4053177_US11153896_257236323181_S01_GE1_107_Sep09_2_1.txt.gz 12.4 Mb (ftp)(http) TXT
Processed data included within Sample table

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