Cytogenetic analysis of interphase cells using Spectral Imaging [SIm] technology

Spectral Imaging (SIm) has dramatically improved our ability to localize and quantitatively analyze multiple nucleic acid targets such as chromosomes, genes and gene transcripts. Studies on metaphase cells such as 'Spectral Karyotyping (SKY)' are less complicated than interphase cell studies because the objects (chromosomes) are spatially separated and ratio-labeled probes can be used to uniquely stain each chromosome type. Our research, however, targets the extensive cytogenetic and phenotypic analysis of interphase cells. The complex organization of interphase chromatin and co-localization of gene transcripts (RNAs) in nuclear or cytoplasmic domains requires unique fluorochrome-labeling for each nucleic acid target. An increasing number of commercially available dyes for probe labeling and software to deconvolute partially overlapping emission spectra has helped to overcome most of these obstacles. This presentation summarizes our experience in analyzing numerical and structural alterations in various human cell types (leukocytes, amniocytes, blastomeres or solid tissue) as well as our approach to multi-gene expression profiling using Sim. Examples illustrate a wide spectrum of groundbreaking techniques for interphase cell analysis. We demonstrate how ten or more chromosomes can be scored in interphase nuclei or the relative level of expression of different transforming RNAs in tumor cells can be measured by SIm.