|
| Status |
Public on May 11, 2017 |
| Title |
Flu_pos_13 |
| Sample type |
RNA |
| |
|
| Source name |
influenza infected patient
|
| Organism |
Homo sapiens |
| Characteristics |
infection: Influenza A
age: 75
tissue: Blood
|
| Extracted molecule |
total RNA |
| Extraction protocol |
A venous blood sample (2.5ml) was collected at the time of enrolment into the study. The sample was placed into PAXgene Blood RNA tubes (BD systems, NJ, USA) and was kept in storage at -20°C. RNA extraction was later performed using the standard protocol (PAXgeneTM Blood RNA kit - Qiagen). RNA was quantified using a NanoDrop-1000 spectrophotometer and quality was monitored with the Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA).
|
| Label |
Cy3
|
| Label protocol |
Cyanine-3 (Cy3) labeled cRNA was prepared from 0.1 ug RNA using the One-Color Low RNA Input Linear Amplification PLUS kit (Agilent) according to the manufacturer's instructions, followed by RNAeasy column purification (QIAGEN, Valencia, CA). Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.
|
| |
|
| Hybridization protocol |
1.5 ug of Cy3-labelled cRNA (specific activity >10.0 pmol Cy3/ug cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 250 ml containing 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturers instructions. On completion of the fragmentation reaction, 250 ml of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to Agilent Whole Human Genome Oligo Microarrays (G4112A) for 17 hours at 65C in a rotating Agilent hybridization oven. After hybridization, microarrays were washed 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute with 37°C GE Wash buffer 2 (Agilent), then dried immediately by brief centrifugation.
|
| Scan protocol |
Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner (G2505B) using one color scan setting for 8x60K array slides (Scan Area 61x21.6 mm, Scan resolution 10um, Dye channel is set to Green and Green PMT is set to 100%).
|
| Data processing |
The scanned images were analyzed with Feature Extraction Software10.5 (Agilent) using default parameters to obtain background subtracted and spatially detrended Processed Signal intensities.
|
| |
|
| Submission date |
May 31, 2016 |
| Last update date |
May 11, 2017 |
| Contact name |
Robert Geffers |
| E-mail(s) |
robert.geffers@helmholtz-hzi.de
|
| Phone |
+49 531-6181-3058
|
| Organization name |
HCI - Helmholtz Centre for Infection Research
|
| Department |
Dep. Molecular Bacteriology
|
| Lab |
Genome Analytics
|
| Street address |
Inhoffenstr. 7
|
| City |
Braunschweig |
| ZIP/Postal code |
38124 |
| Country |
Germany |
| |
|
| Platform ID |
GPL21185 |
| Series (1) |
| GSE82050 |
IFI27 is a gene-expression biomarker that identifies influenza-specific immune response in symptomatic patients |
|
