Mohapatra Gayatry, Betensky Rebecca A, Miller Ezra R, Carey Bjorn, Gaumont Leah D, Engler David A, Louis David N
Department of Pathology, Cancer Center and Neurosurgical Service, Massachusetts General Hospital and Harvard Medical School, Boston 02129, USA.
J Mol Diagn. 2006 May;8(2):268-76. doi: 10.2353/jmoldx.2006.050109.
Array-based comparative genomic hybridization (aCGH) is a powerful, high-throughput tool for whole genome analysis. Until recently, aCGH could only be reproducibly performed on frozen tissue samples and with significant tissue amounts. For brain tumors however, paraffin-embedded tissue blocks from small stereotactic biopsies may be the only tissue routinely available. The development of methods to analyze formalin-fixed, paraffin-embedded (FFPE) material therefore has the potential to impact molecular diagnosis in a significant way. To this end, we constructed a BAC array representing chromosomes 1, 7, 19, and X because 1p/19q deletion and EGFR gene amplification provide clinically relevant information for glioma diagnosis. We also optimized a two-step labeling procedure using an amine-modified nucleotide for generating aCGH probes. Using this approach, we analyzed a series of 28 FFPE oligodendroglial tumors for alterations of chromosomes 1, 7, and 19. We also independently assayed these tumors for 1p/19q deletion by fluorescence in situ hybridization and by loss of heterozygosity analyses. The concordance between aCGH, standard loss of heterozygosity and fluorescence in situ hybridization was nearly 100% for the chromosomes analyzed. These results suggest that aCGH could offer an improved molecular diagnostic approach for gliomas because of its ability to detect clinically relevant molecular alterations in small FFPE specimens.
基于微阵列的比较基因组杂交（aCGH）是一种用于全基因组分析的强大的高通量工具。直到最近，aCGH还只能在冷冻组织样本上且需要大量组织才能可重复地进行。然而，对于脑肿瘤来说，来自小立体定向活检的石蜡包埋组织块可能是唯一常规可用的组织。因此，分析福尔马林固定、石蜡包埋（FFPE）材料的方法的发展有可能对分子诊断产生重大影响。为此，我们构建了一个代表1号、7号、19号染色体和X染色体的BAC微阵列，因为1p/19q缺失和EGFR基因扩增为胶质瘤诊断提供了临床相关信息。我们还优化了一种使用胺修饰核苷酸生成aCGH探针的两步标记程序。使用这种方法，我们分析了一系列28个FFPE少突胶质细胞瘤的1号、7号和19号染色体的改变。我们还通过荧光原位杂交和杂合性缺失分析独立检测了这些肿瘤的1p/19q缺失情况。对于所分析的染色体，aCGH、标准杂合性缺失和荧光原位杂交之间的一致性接近100%。这些结果表明，aCGH因其能够在小的FFPE样本中检测到临床相关的分子改变，可为胶质瘤提供一种改进的分子诊断方法。
