|
| Status |
Public on Feb 01, 2010 |
| Title |
H2O 3d 4 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
HaCaT cells pool
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: keratinocytes (HaCaT)
reference: common reference
|
| Treatment protocol |
Cells were exposed to 30 µg/ml WC (153 µM), 33 µg/ml WC-Co (153/51 µM), or 3 µg/ml (51 µM) cobalt chloride by mixing RPMI 5 % FBS with 10 fold concentrated stock solutions. Exposure was performed in the dark with 5 independent replicates (performed at different days using different cell passage numbers). Controls were performed by with the water used for the preparation of particle suspensions.
|
| Growth protocol |
HaCaT cells were maintained in RPMI medium (‘Roswell Park Memorial Institute’ medium; Biochrom, Karlsruhe, Germany) supplemented with 5 % (v/v) FBS and 1% (v/v) penicillin/streptomycin. Cells were cultured in monolayers at 37°C in a humidified, 5 % (v/v) CO2-atmosphere and sub-cultured twice a week in 75 cm² flasks. Cells were seeded at densities of 200.000 cells/ml for 3d of exposure or 500.000 cells/ml for 3h of exposure, respectively in a final volume of 10 ml per 75 cm² flasks.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA extracted using Trizol following manufacturer's instructions.
|
| Label |
Cy5
|
| Label protocol |
Synthesis of cDNA, cRNA and cRNA-labeling was performed with the Agilent Low RNA Input Linear Amplification Kit according to the manufacturer’s instructions. cRNA was labelled with Cy3 (controls and treatments) and Cy5 (common reference).
|
| |
|
| Channel 2 |
| Source name |
HaCaT cells
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: keratinocytes (HaCaT)
exposure time: 3 days
treatment: H2O
|
| Treatment protocol |
Cells were exposed to 30 µg/ml WC (153 µM), 33 µg/ml WC-Co (153/51 µM), or 3 µg/ml (51 µM) cobalt chloride by mixing RPMI 5 % FBS with 10 fold concentrated stock solutions. Exposure was performed in the dark with 5 independent replicates (performed at different days using different cell passage numbers). Controls were performed by with the water used for the preparation of particle suspensions.
|
| Growth protocol |
HaCaT cells were maintained in RPMI medium (‘Roswell Park Memorial Institute’ medium; Biochrom, Karlsruhe, Germany) supplemented with 5 % (v/v) FBS and 1% (v/v) penicillin/streptomycin. Cells were cultured in monolayers at 37°C in a humidified, 5 % (v/v) CO2-atmosphere and sub-cultured twice a week in 75 cm² flasks. Cells were seeded at densities of 200.000 cells/ml for 3d of exposure or 500.000 cells/ml for 3h of exposure, respectively in a final volume of 10 ml per 75 cm² flasks.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA extracted using Trizol following manufacturer's instructions.
|
| Label |
Cy3
|
| Label protocol |
Synthesis of cDNA, cRNA and cRNA-labeling was performed with the Agilent Low RNA Input Linear Amplification Kit according to the manufacturer’s instructions. cRNA was labelled with Cy3 (controls and treatments) and Cy5 (common reference).
|
| |
|
| |
| Hybridization protocol |
Hybridization was performed using the Agilent Gene Expression Hybridization Kit following manufacturer's instructions.
|
| Scan protocol |
Scanned on an Agilent scanner. Images were quantified using Agilent Feature Extraction Software (version 9.5.3.1 or 10.1.1.1).
|
| Description |
Biological replicate 3 of 5, harvested after several passages.
|
| Data processing |
Feature extraction files (processed signals) were used for statistics.
|
| |
|
| Submission date |
Jun 19, 2009 |
| Last update date |
Dec 22, 2009 |
| Contact name |
Wibke Busch |
| E-mail(s) |
[email protected]
|
| Phone |
00493412351515
|
| Organization name |
UFZ Helmholtz Centre for Environmental Research
|
| Department |
Bioanalytical Ecotoxicology
|
| Lab |
Scholz
|
| Street address |
Permoser Str. 15
|
| City |
Leipzig |
| ZIP/Postal code |
04318 |
| Country |
Germany |
| |
|
| Platform ID |
GPL4133 |
| Series (1) |
| GSE16727 |
Human keratinocytes exposed to tungsten carbide and tungsten carbide cobalt nanoparticles and cobalt chloride |
|
