Microarray-Based Comparative Genomic Hybridization

Comparative genomic hybridization based on microarrays was performed in a commercial whole-genome 180 K platform containing 180,000 oligonucleotide probes (Agilent Technologies; design 22060), using DNA from the tumor sample. Reference DNA was a commercially available human pool of samples from multiple anonymous healthy donors (Promega Corporation). Technical procedures are described in Torrezan et al. [11 (link)]. Hybridization and washing were performed as recommended by the manufacturer. Scanned images were processed using Feature Extraction 10.7.3.1 software (Agilent Technologies), and array CGH analysis was conducted with Nexus Copy Number software 7.0 (Biodiscovery). We used the FASST2 segmentation algorithm, according to the following settings: minimum of five consecutive probes (effective resolution of ~70 Kb for CNA calling), significance threshold set at 10⁻⁸, and threshold log₂ Cy3/Cy5 of 0.33 and −0.3 for gains for loss, respectively, and 1.2 and −1.1 for high copy number gains and homozygous losses, respectively. All copy number alterations are reported in the Database of Genomic Variants [12 ].