Department of Obstetrics, Gynecology and Reproductive Sciences, University of California, San Francisco, California.
Life Sciences Division, E.O. Lawrence Berkeley National Laboratory, Berkeley, California.
J Histochem Cytochem. 2018 Aug;66(8):595-606. doi: 10.1369/0022155418771613. Epub 2018 Apr 19.
Aneuploidy seems to play not only a decisive role in embryonal development but also in tumorigenesis where chromosomal and genomic instability reflect a universal feature of malignant tumors. The cost of whole genome sequencing has fallen significantly, but it is still prohibitive for many institutions and clinical settings. No applied, cost-effective, and efficient technique has been introduced yet aiming at research to assess the ploidy status of all 24 different human chromosomes in interphases simultaneously, especially in single cells. Here, we present the selection of human probe DNA and a technique using multistep fluorescence in situ hybridization (FISH) employing four sets of six labeled FISH probes able to delineate all 24 human chromosomes in interphase cells. This full karyotype analysis approach will provide additional diagnostic potential for single cell analysis. The use of spectral imaging (SIm) has enabled the use of up to eight different fluorochrome labels simultaneously. Thus, scoring can be easily assessed by visual inspection, because SIm permits computer-assigned and distinguishable pseudo-colors to each probe during image processing. This enables full karyotype analysis by FISH of single-cell interphase nuclei.
非整倍体似乎不仅在胚胎发育中起决定性作用,而且在肿瘤发生中也起作用,其中染色体和基因组不稳定性反映了恶性肿瘤的普遍特征。全基因组测序的成本已经大幅下降,但对许多机构和临床环境来说仍然过高。目前还没有引入任何实用、经济高效的技术,旨在研究同时评估所有 24 个人类染色体的倍性状态,特别是在单细胞中。在这里,我们选择了人类探针 DNA,并使用多步荧光原位杂交 (FISH) 技术,采用四组六个标记的 FISH 探针,能够在间期细胞中描绘所有 24 个人类染色体。这种全染色体组分析方法将为单细胞分析提供额外的诊断潜力。光谱成像 (SIm) 的使用使得可以同时使用多达八个不同的荧光染料标签。因此,通过视觉检查可以轻松评估评分,因为 SIm 在图像处理过程中允许为每个探针分配可区分的伪彩色。这使得通过 FISH 对单细胞间期核进行全染色体组分析成为可能。