|
| Status |
Public on Aug 01, 2019 |
| Title |
B16F0 cells cultured for 24 hr in serum-free media, rep 2 |
| Sample type |
RNA |
| |
|
| Source name |
B16F0 cells
|
| Organism |
Mus musculus |
| Characteristics |
tag: B16F0
|
| Treatment protocol |
not applicable
|
| Growth protocol |
Cultured for 24 hours in serum-free media
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA isolated from B16F0 exosomes and cells by RNeasy kit (Qiagen) was quantified using Nanodrop and analyzed by on-chip-electrophoresis using the Agilent Bioanalyzer.
|
| Label |
biotin
|
| Label protocol |
Four hundred nanograms of each miRNA sample was biotin labeled using the Genisphere FlashTag HSR Kit according to the manufacturer's instructions. The labeling of RNA was confirmed by Enzyme Linked Oligosorbent Assay (ELOSA) according Genisphere FlashTag HSR protocol.
|
| |
|
| Hybridization protocol |
The 21.5 microl of biotin-labeled RNA with added hybridization controls was hybridized to the GeneChip miRNA 2.0 Arrays (Affymetrix) at 48oC and 60 rpm for 16 hours in GeneChip Hybridization Oven 640 (Affymetrix).The entire reaction of fragmented and biotin-labeled cDNA (50 mu) with added hybridization controls was hybridized to the mouse GeneChip 1.0 ST Exon Arrays (Affymetrix) at 45oC for 17 hours in GeneChip Hybridization Oven 640 (Affymetrix).
|
| Scan protocol |
GeneChip miRNA 2.0 Arrays were stained using FS 450_0003 protocol in Affymetrix GeneChip Fluidics Station 450. Briefly, biotin-labeled RNA was reacted using washes with a solution containing a streptavidin-phycoerythrin complex, with an intermediate treatment of biotin-labeled anti-streptadvidin antibody to amplify the signal. Phycoerythrin labeling was detected within the Affymetrix GeneChip Scanner 3000 7G plus using 532 nm light and detected by a photomultiplier tube.
|
| Data processing |
A miRNA QC Tool software (Affymetrix) was used to check quality controls of hybridized chips. All chips that passed quality controls were RMA normalized using miRNA QC Tool.
|
| |
|
| Submission date |
Aug 21, 2017 |
| Last update date |
Aug 01, 2019 |
| Contact name |
David John Klinke |
| E-mail(s) |
David.Klinke@mail.wvu.edu
|
| Phone |
304-293-9346
|
| Organization name |
West Virginia University
|
| Department |
Chemical Engineering
|
| Street address |
P.O. Box 6102
|
| City |
Morgantown |
| State/province |
WV |
| ZIP/Postal code |
26506-6102 |
| Country |
USA |
| |
|
| Platform ID |
GPL14613 |
| Series (2) |
| GSE102883 |
Expression data of miRNA from parental B16F0 cells and B16F0 exosomes [miRNA] |
| GSE102951 |
Expression data from parental B16F0 cells and B16F0 exosomes |
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