MOLT4 cells were exposed to a increased concentrations of MTX, until final concentrations of 300 nM, respectively, were reachedFor this purpose the cells were cultured and sub-cultured bi- weekly in RPMI-1640 medium containing fetal calf serum (10%), penicillin (100 U/ml), streptomycin (100 µg/ml) and L-glutamine (2 mM) and under humidified air containing 5% CO2 and at 37?C. The cultures were tested routinely for contamination by mycoplasma. The cells were cultured in drug-free-medium for at least 3 passages prior to harvesting during logarithmic phase growth (at a density of approximately 0.8-1.5x106/ml) for experimental studies. A Coulter Multi-sizer (Coulter Electronics, Luton, United Kingdom) was used to count the cells.
Extracted molecule
total RNA
Extraction protocol
RNeasy Midi kit in accordance with manufacturer’s instructions
Label
biotin
Label protocol
Biotinylated cRNA were prepared according to the standard Affymetrix protocol from 6 microg total RNA (Expression Analysis Technical Manual, 2001, Affymetrix).
Hybridization protocol
Following fragmentation, 10 microg of cRNA were hybridized for 16 hr at 45C on GeneChip human U133A Genome Array. GeneChips were washed and stained in the Affymetrix Fluidics Station 400.
Scan protocol
GeneChips were scanned using the Hewlett-Packard GeneArray Scanner G2500A.
Description
Gene expression data from untreated MOLT4 cells
Data processing
The data were analyzed with Microarray Suite version 5.0 (MAS 5.0) using Affymetrix default analysis settings and global scaling as normalization method. The trimmed mean target intensity of each array was arbitrarily set to 100.
