A Genome-Wide High-Resolution Array-CGH Analysis of Cutaneous Melanoma and Comparison of Array-CGH to FISH in Diagnostic Evaluation  Lu Wang, Mamta Rao, Yuqiang Fang, Meera Hameed, Agnes Viale, Klaus Busam, Suresh C. Jhanwar  The Journal of Molecular Diagnostics  Volume 15, Issue 5, Pages 581-591 (September 2013) DOI: 10.1016/j.jmoldx.2013.04.001 Copyright © 2013 American Society for Investigative Pathology and the Association for Molecular Pathology Terms and Conditions

Figure 1 Summary of workflow for aCGH data analysis, process, and interpretation. CNA, copy number aberration; CNV, copy number variation; DGV, Database of Genomic Variants (http://dgv.tcag.ca/dgv); TSG, tumor suppressor gene. The Journal of Molecular Diagnostics 2013 15, 581-591DOI: (10.1016/j.jmoldx.2013.04.001) Copyright © 2013 American Society for Investigative Pathology and the Association for Molecular Pathology Terms and Conditions

Figure 2 Aggregated copy number aberration in 25 melanoma samples identified via aCGH 244K in the present study. In the genome view, blue represents copy number gain and red represents copy number loss. Dotted lines on the right side of each chromosome idiogram indicate the frequency (0% to 100%) of the identified aberrations. For example, loss of chromosome 9p was identified in >40% of melanoma tumor samples. In addition, deletion of CDKN2A (indicated by the arrow) was detected in 52% of melanoma samples. The Journal of Molecular Diagnostics 2013 15, 581-591DOI: (10.1016/j.jmoldx.2013.04.001) Copyright © 2013 American Society for Investigative Pathology and the Association for Molecular Pathology Terms and Conditions

Figure 3 Comparison of aCGH and FISH assays in evaluation of copy number change in the CDKN2A gene in five melanoma samples. For each case except Mela1, a schematic DNA copy number profile across chromosome 9 revealed via aCGH 244K analysis and a representative FISH image of the CDKN2A/CEP9 probe set are shown side by side. For Mela12, a representative H&E-stained morphologic image is shown next to its FISH image. Red arrows indicate the locus of CDKN2A. For FISH, the CDKN2A gene is labeled in red, and CEP9 in green. For Mela1, no specimen was available for FISH assay. For Mela5, homozygous deletion of CDKN2A (a deletion segment of 428 kb) and hemizygous deletion of the remainder of chromosome 9p was detected via aCGH, and FISH confirmed homozygous deletion of CDKN2A. For Mela20, homozygous deletion of CDKN2A (a deletion segment of 93 Kb) and hemizygous deletion of the remainder of chromosome 9p was detected via aCGH; at FISH, one signal for CDKN2A was present but was unusually smaller than commonly observed, despite the expected hybridization quality. The FISH probe was about 190 kb, twice the size of the deletion segment detected via aCGH. Therefore, the unusually small signal was considered to be that of the residual DNA sequence. For Mela24, homozygous deletion of CDKN2A (a deletion segment of 68 kb) and hemizygous deletion of the remainder of chromosome 9p was detected via aCGH; at FISH, as for Mela20, one signal for CDKN2A was present but was unusually smaller than commonly observed, despite the expected hybridization quality. For Mela12, a 554-kb hemizygous deletion on 9p21.3 overlapping the entire CDKN2A gene was detected via aCGH; at FISH, homozygous deletion of CDKN2A was revealed in some (usually large) cells, whereas two signals for CDKN2A were observed in the other (usually small) cells. H&E staining of the same tissue showed melanoma tumor cells in a background of a substantial number of stromal and inflammatory cells. This example shows that aCGH cannot differentiate heterozygous deletion from mosaic tissue with both homozygous deletion and normal populations. The Journal of Molecular Diagnostics 2013 15, 581-591DOI: (10.1016/j.jmoldx.2013.04.001) Copyright © 2013 American Society for Investigative Pathology and the Association for Molecular Pathology Terms and Conditions