|
| Status |
Public on Nov 02, 2011 |
| Title |
human lung adenocarcinoma cell line a - non-permissive for MVwt - cultured - biol rep 2 |
| Sample type |
RNA |
| |
|
| Source name |
original cell source: ATCC (#CRL-5800) - normal culture under optimal culture conditions -> cells in exponential growth phase
|
| Organism |
Homo sapiens |
| Characteristics |
disease: lung adenocarcinoma
cell line: CRL-5800
virus permissive: no
|
| Growth protocol |
Cell lines were grown in T175 flasks until sub-confluence under optimal growth conditions. The cells were detached from the culture vessel using PBS-EDTA (1 mM) + Trypsin, and pelleted at 500xg for 5 min at 4°C. Supernatants were removed, the cells were washed once with 1 ml ice-cold PBS and pelleted, again. Supernatants were aspirated and the cell pellets were snap-frozen by submersion in liquid nitrogen for approx. 1 min. Frozen cell pellets were stored at -80°C until shipment of samples for extraction and further analysis on dry ice.
Cell lines were cultured as instructed by ATCC. In brief: H23, H358, H441, and H522 were grown in RPMI 1640 + 10% FCS; HT1376 were grown in EMEM + 10% FCS; T24 were grown in McCoy´s 5a + 10% FCS; SCaBER were grwon in MEM (adjusted to contain 1.5 g/L sodium bicarbonate and 0.1 mM non-essentail amino acids) + 2 mM L-Gln + 10% FCS. All cell lines were grown at 37°C in a humidified atmosphere containing 5% carbon dioxide.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was isolated using the NucleoSpin® RNA II kit (Macherey-Nagel) according to the manufacturer's instructions.
|
| Label |
Cy3
|
| Label protocol |
1 µg of each RNA was used as template to produce Cy3- labeled cRNA. The RNA samples were amplified and labeled using the Agilent Low RNA Input Linear Amp Kit (Agilent Technologies) following the manufacturer’s protocol.
|
| |
|
| Hybridization protocol |
Cy3 labeled cRNAs were hybridized overnight (17 hours, 65°C) to Agilent Whole Human Genome Oligo Microarrays (4x44K) using Agilent’s recommended hybridization chamber and oven. Finally, the microarrays were washed once with the Agilent Gene Expression Wash Buffer 1 for 1 min at room temperature followed by a second wash with preheated Agilent Gene Expression Wash Buffer 2 (37 °C) for 1 min. The last washing step was performed with acetonitrile.
|
| Scan protocol |
Fluorescence signals of the hybridized Agilent Microarrays were detected using Agilent’s Microarray Scanner System (G2505C, Agilent Technologies, Palo Alto, USA).
|
| Description |
H23
|
| Data processing |
Feature Extraction Software derived output data files were further processed with Rosetta Resolver (v7.2); Rosetta Inpharmatics LLC
|
| |
|
| Submission date |
Sep 15, 2011 |
| Last update date |
Nov 02, 2011 |
| Contact name |
Michael Mühlebach |
| E-mail(s) |
Michael.Muehlebach@pei.de
|
| Phone |
+49 - 6103 - 77 4005
|
| Fax |
+49 - 6103 - 77 1255
|
| Organization name |
Paul-Ehrlich-Institut
|
| Street address |
Paul-Ehrlich-Strasse 51-59
|
| City |
Langen |
| ZIP/Postal code |
63225 |
| Country |
Germany |
| |
|
| Platform ID |
GPL6480 |
| Series (1) |
| GSE32155 |
Identification of the epithelial receptor for measles virus |
|
