|
| Status |
Public on May 22, 2009 |
| Title |
ACHN_IFN_12hr |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
ACHN cells, untreated, 12 hours
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: Renal Cell Carcinoma immortalized
|
| Treatment protocol |
IFNalpha-2b (Intron A, Schering-Plough, Kenilworth, NJ) was added to fresh medium at a final concentration of 1000 IU/mL. Cells were treated for 3, 6, 9, 12, 16, or 24 hours.
|
| Growth protocol |
RCC1 cells were maintained in RPMI 1640 supplemented with 10 g/mL penicillin and streptomycin and 10% fetal bovine serum (FBS). ACHN (ATCC CRL-1611) cells were cultured in DMEM with 10 g/mL penicillin and streptomycin and 10% FBS.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was harvested post-treatment using Trizol reagent according to the manufacturer’s instructions (Invitrogen, Carlsbad, CA).
|
| Label |
Cy3
|
| Label protocol |
Total RNA (20 g) was used to synthesize first-strand cDNA, which contains aminoallyl dUTP using random priming. The aa dUTP containing cDNA was conjugated to Cy3 (untreated) or Cy5 (treated) ester dye under basic conditions, and free nucleotide was removed by purifying the labeled cDNA with a GFX column (Amersham, Arlington Heights, IL).
|
| |
|
| Channel 2 |
| Source name |
ACHN cells, IFN treated, 12 hours
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: Renal Cell Carcinoma immortalized
|
| Treatment protocol |
IFNalpha-2b (Intron A, Schering-Plough, Kenilworth, NJ) was added to fresh medium at a final concentration of 1000 IU/mL. Cells were treated for 3, 6, 9, 12, 16, or 24 hours.
|
| Growth protocol |
RCC1 cells were maintained in RPMI 1640 supplemented with 10 g/mL penicillin and streptomycin and 10% fetal bovine serum (FBS). ACHN (ATCC CRL-1611) cells were cultured in DMEM with 10 g/mL penicillin and streptomycin and 10% FBS.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was harvested post-treatment using Trizol reagent according to the manufacturer’s instructions (Invitrogen, Carlsbad, CA).
|
| Label |
Cy5
|
| Label protocol |
Total RNA (20 g) was used to synthesize first-strand cDNA, which contains aminoallyl dUTP using random priming. The aa dUTP containing cDNA was conjugated to Cy3 (untreated) or Cy5 (treated) ester dye under basic conditions, and free nucleotide was removed by purifying the labeled cDNA with a GFX column (Amersham, Arlington Heights, IL).
|
| |
|
| |
| Hybridization protocol |
Hybridization was performed in slide hyb 3 hybridization buffer (Ambion, Austin, TX) beneath lifter slip coverslips (Erie Scientific, Portsmouth, NH) at 55°C for 16 h.
|
| Scan protocol |
Slides were washed for 5 min in each of three wash solutions containing 2x SSC/0.1% SDS, 2x SSC, and 0.2x SSC, centrifuged, and scanned using a GenePix 4000 scanner (Axon, Union City, CA).
|
| Description |
081402_ACHN_12 hr_IFN_0001.gpr
|
| Data processing |
Array spots were defined using GenePix software and data were analyzed using GeneSpring (Silicon Genetics, Agilent Technologies, Palo Alto, CA). SNOMAD was used for normalization and background correction (pevsnerlab.kennedykrieger.org/snomadinput.html). Expression Analysis Systematic Explorer (EASE) was used to identify functional similarities within gene clusters (david.niaid.nih.gov/david/ease.htm).
|
| |
|
| Submission date |
Mar 11, 2009 |
| Last update date |
May 22, 2009 |
| Contact name |
Michelle Holko |
| E-mail(s) |
holko@ncbi.nlm.nih.gov
|
| Phone |
(301) 496-5753
|
| Organization name |
GEO
|
| Department |
NCBI
|
| Street address |
45 Center Dr
|
| City |
Bethesda |
| State/province |
MD |
| ZIP/Postal code |
20892 |
| Country |
USA |
| |
|
| Platform ID |
GPL8281 |
| Series (2) |
| GSE15193 |
IFN-alpha Stimulated Gene Expression in Sensitive and Resistant Renal Cell Carcinoma Cell Lines |
| GSE16197 |
Transcription Factor Promyelocytic Leukemia Zinc Finger Protein regulates Interferon Stimulated Gene expression |
|
