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Sample GSM4053173 Query DataSets for GSM4053173
Status Public on Sep 04, 2020
Title Donor 1, Timepoint 0, Replicate A2
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 0)
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
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
VALUE quantile

Data table
1 16.80637437
2 4.500387398
3 4.682805067
4 5.407690902
5 4.626990545
6 5.312099863
7 5.951851555
8 12.56818627
9 4.795597271
10 5.142550934
11 4.848533214
12 4.848533214
13 5.389726221
14 4.82263073
15 11.69626628
16 4.90081848
17 6.534348876
18 10.62726311
19 5.270597986
20 6.649787508

Total number of rows: 62976

Table truncated, full table size 1086 Kbytes.

Supplementary file Size Download File type/resource
GSM4053173_US11153896_257236323182_S01_GE1_107_Sep09_1_1.txt.gz 12.3 Mb (ftp)(http) TXT
Processed data included within Sample table

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