Kallioniemi O P, Kallioniemi A, Piper J, Isola J, Waldman F M, Gray J W, Pinkel D
Department of Laboratory Medicine, Tampere University Hospital.
Genes Chromosomes Cancer. 1994 Aug;10(4):231-43. doi: 10.1002/gcc.2870100403.
Comparative genomic hybridization (CGH) is a powerful new method for molecular cytogenetic analysis of cancer. In a single hybridization, CGH provides an overview of DNA sequence copy number changes (losses, deletions, gains, amplifications) in a tumor specimen and maps these changes on normal chromosomes. CGH is based on the in situ hybridization of differentially labeled total genomic tumor DNA and normal reference DNA to normal human metaphase chromosomes. After hybridization and fluorescent staining of the bound DNAs, copy number variations among the different sequences in the tumor DNA are detected by measuring the tumor/normal fluorescence intensity ratio for each locus in the target metaphase chromosomes. CGH is in particular useful for analysis of DNA sequence copy number changes in common solid tumors where high-quality metaphase preparations are often difficult to make, and where complex karyotypes with numerous markers, double minutes, and homogeneously stained chromosomal regions are common. CGH only detects changes that are present in a substantial proportion of tumor cells (i.e., clonal aberrations). It does not reveal translocations, inversions, and other aberrations that do not change copy number. At present, CGH is a research tool that complements previous methods for genetic analysis. CGH will advance our understanding of the genetic progression of cancer and highlight important genomic regions for further study. Direct clinical applications of CGH are possible, but will require further development and validation of the technique. We describe here our recent optimized procedures for CGH, including DNA labeling, hybridization, fluorescence microscopy, digital image analysis, data interpretation, and quality control, emphasizing those steps that are most critical. We will also assess sensitivity and resolution limits of CGH as well as discuss possible future technical improvements.
比较基因组杂交（CGH）是一种用于癌症分子细胞遗传学分析的强大新方法。在一次杂交中，CGH可提供肿瘤标本中DNA序列拷贝数变化（缺失、删除、增加、扩增）的概况，并将这些变化定位到正常染色体上。CGH基于差异标记的肿瘤全基因组DNA和正常参照DNA与正常人中期染色体的原位杂交。在对结合的DNA进行杂交和荧光染色后，通过测量目标中期染色体上每个位点的肿瘤/正常荧光强度比，检测肿瘤DNA中不同序列之间的拷贝数变异。CGH对于分析常见实体瘤中的DNA序列拷贝数变化特别有用，因为在这些肿瘤中高质量的中期染色体标本往往难以制备，并且具有众多标记、双微体和均匀染色染色体区域的复杂核型很常见。CGH仅检测存在于相当比例肿瘤细胞中的变化（即克隆性畸变）。它不会揭示不改变拷贝数的易位、倒位和其他畸变。目前，CGH是一种补充先前遗传分析方法的研究工具。CGH将增进我们对癌症遗传进展的理解，并突出重要的基因组区域以供进一步研究。CGH的直接临床应用是可能的，但需要对该技术进行进一步的开发和验证。我们在此描述我们最近优化的CGH程序，包括DNA标记、杂交、荧光显微镜检查、数字图像分析、数据解释和质量控制，重点强调那些最关键的步骤。我们还将评估CGH的灵敏度和分辨率极限，并讨论未来可能的技术改进。
