Briefly, whole cell lysates were prepared from Selleck IBET762 microdissected areas containing 70% tumors. DNA from lysates was processed in parallel with matched CSFTCs, as outlined in the next section. Array comparative genomic hybridization (ACGH) analysis usually requires DNA input that is roughly equivalent to genomic material from several thousands of cells. Since CSFTCs are rare, we can only isolate small pools of CSFTCs. The few hundred picograms of genomic material from these few cells require whole genome amplification (WGA) prior to downstream molecular analysis. To reduce the likelihood of detecting false positives (due to amplification bias) when comparing CSFTCs versus matched archival tumors, we subjected both sets of tumors to the same WGA method [4]. Samples from 2 of the 15 patients failed WGA product quality testing [4] and were excluded from further analysis. Amplified tumor DNA samples were then subjected to ACGH analysis using a bacterial artificial chromosome (BAC) array containing 2464 clones printed in triplicate [5]. The BAC arrays were printed at the UCSF Helen Diller Family Comprehensive Cancer Center Array Core. The ACGH experimental protocol has been previously described in detail [4]. Briefly, the tumor (test sample) and reference DNA samples were differentially labeled with Cy3 and Cy5 dyes, respectively, and co-hybridized to a BAC array. A sex-mismatched (i.e., female vs. male) hybridization was used as an internal control to detect a copy number gain in X- and loss of Y-chromosomes in the female test sample. Post-hybridization imaging data and analysis of the BAC array were done as previously described [6]. The intensity values were used to calculate Cy3/Cy5 ratios using the UCSF Spot Program. An in-house R package ‘Spot Correction’ was also used to remove systematic variations of unknown origin across the array [7], including a correction that is based on the GC content of the BAC clones [4]. The aCGH data was processed using the custom program SPROC [8] in order to automatically filter out data points with low DAPI intensity, low correlation between Cy3 and Cy5 within each spot, and low reference/DAPI signal intensity. Clones whose ratios that were derived from only one of the triplicate spots or with a triplicate log2 SD > 0.2 were set as “missing”. The clones were mapped to the May 2004 freeze of the human DNA sequence. The median absolute deviation (MAD) estimates (see below) were used as a measure of the quality of the microarray data. Array data with a MAD estimate < 0.25 was considered “good quality”. A histogram of MAD estimates showed that all samples passed the MAD threshold (Fig. 1). Visual inspection of each aCGH profile was also performed to confirm results. The microarray and sample annotation data for 30 samples from 13 patients were deposited in Gene expression Omnibus under accession number GSE46068.