|
| Status |
Public on Dec 15, 2012 |
| Title |
SAE_MWCNT_100_1hr-2 |
| Sample type |
RNA |
| |
|
| Source name |
SAE cells exposed to 100 ug/ml MWCNT for 1hr, biological rep2
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: SAE
cell type: small airway epithelial
treatment: MWCNT
dose: 100 ug/ml
time: 1 hr
|
| Treatment protocol |
For transcriptomics analysis, cells were suspended at 1 x 106 cells per ml and plated in 25 cm2 flasks at 5 x 106 cells/flask. The multi-walled carbon nanotubes (MWCNT) or titanium nanobelts (TiO2-NB) were added directly to the wells at specified concentrations (0, 10 or 100 µg/ml) in media. The cells were cultured for either 1 or 24 h. At the end of the culture period the adherent cells were washed once in PBS and the RNA was isolated as described below.
|
| Growth protocol |
Small airway epithelial (SAE) cells were obtained from Lonza Walkersville Inc. Cells were maintained in Lonza Clonetics SABM media (cat # cc-3119) supplemented with Lonza Clonetics SAGM single quots (cat# cc-4124). For experimental conditions, adherent SAEC cells were washed with HBSS (cat# cc-5022) and trypsin (cat# cc-5012) was then used to dislodge the cells. A trypsin neutralizing solution (cat# cc-5002) was then added and the cells were centrifuged at 250 x g for 5 min. The supernatant was discarded and the cells were resuspended and counted as stated above. All cultures were maintained in 37°C water-jacketed CO2 incubators (ThermoForma).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was collected using the RNeasy Kit (Qiagen, Valencia, CA).
|
| Label |
biotin
|
| Label protocol |
Complementary DNA was synthesized from 3 µg of total RNA in the presence of an oligo-dT primer containing a T7 RNA polymerase promoter, and an in vitro transcription reaction was performed in the presence of a mixture of biotin-labeled ribonucleotides to produce biotinylated cRNA from the cDNA template, according to manufacturer’s protocols (Affymetrix One-Cycle Target Labeling Kit).
|
| |
|
| Hybridization protocol |
Biotin-labeled cRNA (15 µg) was fragmented to a size range between 50-200 bases for array hybridization. After hybridization, the arrays were washed and stained with streptavidin-phycoerythrin.
|
| Scan protocol |
The arrays were scanned at a resolution of 2.5 microns using an Affymetrix GeneChip Scanner 3000.
|
| Description |
Gene expression data from SAE cells exposed to 100 ug/ml MWCNT for 1hr
|
| Data processing |
Raw intensity data were quantile normalized by Robust Multi-Array Analysis (RMA) summarization and probes were subject to quality control to measure the efficiency of transcription, integrity of hybridization, and consistency of qualitative calls.
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| |
|
| Submission date |
Nov 06, 2012 |
| Last update date |
Dec 15, 2012 |
| Contact name |
Susan C. Tilton |
| E-mail(s) |
Susan.Tilton@oregonstate.edu
|
| Organization name |
Oregon State University
|
| Department |
AG Envr & Molec Toxicology
|
| Street address |
1007 ALS Bldg
|
| City |
Corvallis |
| State/province |
OR |
| ZIP/Postal code |
97331 |
| Country |
USA |
| |
|
| Platform ID |
GPL571 |
| Series (2) |
| GSE42067 |
Three human cell types respond to multi-walled carbon nanotubes and titanium dioxide nanobelts with cell-specific transcriptomic and proteomic expression [SAE cells] |
| GSE42069 |
Three human cell types respond to multi-walled carbon nanotubes and titanium dioxide nanobelts with cell-specific transcriptomic and proteomic expression |
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