Human Genome CGH Microarray 244A Supplemental

Protocol tips

Upstream tips	Protocol tips	Downstream tips
Avoid touching the surface of the glass with your fingers. Skin oils and other substances, such as lotions or ink, can fluoresce. If the surface of the glass is noticeably dirty, it can be carefully cleaned with a nonabrasive laboratory tissue.

Upstream tips
Avoid touching the surface of the glass with your fingers. Skin oils and other substances, such as lotions or ink, can fluoresce. If the surface of the glass is noticeably dirty, it can be carefully cleaned with a nonabrasive laboratory tissue.

Publication protocol

CGH analysis
Genomic DNA (2 μg) isolated from the tissue samples and DNA from healthy tissue (Promega Corporation, Madison, WI) were labeled with Cy5 and Cy3, respectively, using an Oligo aCGH Labeling Kit (Agilent Technologies). The Cy5-labeled and Cy3-labeled samples were purified using Agilent-KREApure columns (Agilent Technologies). The appropriately purified and labeled samples were processed according to the manufacturer’s protocol. Before hybridization to the array, the hybridization mixtures were denatured at 95 °C for 3 min and incubated at 37 °C for 30 min, and the samples were then applied to the Agilent Genome CGH Microarray 244 K (Agilent Technologies). Hybridization was carried out for 40 h at 65 °C. After washing, the slides were scanned on an Agilent microarray scanner. The images were analyzed using Feature Extraction software, version 10.5.1.1 (Agilent Technologies) and Agilent Genomic Workbench software, version 7.0.4.0 (Agilent Technologies) using the ADM-2 algorithm with the following settings: 2 Mb moving average; threshold, 6; filter, 3 probes; and 0.4 log2 ratio.

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