Non-Biological Variability Study

Assessment of sources of non-biological (technical) variability in the processing of RNA samples and gene chips during microarray analysis. The purpose of this experiment was to measure technical variability in microarray data introduced by various steps in preparation of the RNA, amplification/labeling of the RNA and hybridization of the sample to the arrays on separate days. The experimental design utilized a single tissue source - homogenized brain tissue pooled from 3 eight week old male C57/Bl mice. Variables examined include: I) Technician extracting RNA (technician #1 prepared Pools 1 and 2; technician #2 prepared pools 3 and 4). II) RNA extraction (Pools 1-4 represent four independent RNA extractions from 3 homogenized pooled whole mouse brains from 8 week old male C57/B6 mice) III) Independent RNA labeling/amplification reactions - three performed from each RNA pool labeled A, B and C) and IV) Day of hybridization to GLYCOv1 arrays (labeled samples were hybridized to arrays over 3 days - 11/14/02, 11/15/02 and 11/18/02). 32 samples made it through to completion generating a high quality .CEL file for analysis. Four samples failed at various steps in the preparation/hybridiztion. All samples were male 8 week old C57/Bl mouse brain total RNA.