Repeated DNAs from the constitutive heterochromatin of human chromosomes 1 and 18 were used as probes in nonradioactive in situ hybridization experiments to define specific numerical and structural chromosome aberrations in three human glioma cell lines and one neuroblastoma cell line. The number of spots detected in interphase nuclei of these tumor cell lines and in normal diploid nuclei correlated well with metaphase counts of chromosomes specifically labeled by in situ hybridization. Rapid and reliable assessments of aneuploid chromosome numbers in tumor lines in double hybridization experiments were achieved, and rare cells with bizarre phenotype and chromosome constitution could be evaluated in a given tumor cell population. Even with suboptimal or rare chromosome spreads specific chromosome aberrations were delineated. As more extensive probe sets become available this approach will become increasingly powerful for uncovering various genetic alterations and their progression in tumor cells. 0 1988 Academic Press. Inc.Specific chromosome aberrations are thought to be important in the development of malignant human gliomas. However, several groups have noted that adequate chromosome analyses of both primary cultures and of solid glioma samples are problematic [ 1, 21. These difficulties are generally encountered in all cytogenetic analyses of solid tumors; many tumor preparations do not contain sufficient numbers of well-spread and banded metaphase cells, and a large number of mitotic cells within solid tumors are inaccessible for analysis. With such small samples of mitotic cells it is difficult to determine if a chromosomal pattern is representative of the whole tumor cell population. Thus specific numerical and structural aberrations in a given tumor type are difficult to confirm on a broad scale, and rare aberrant cells with special malignant propensities may also be overlooked. Furthermore the analysis of complex chromosome changes in very aneuploid cells requires the detailed attention of highly skilled observers. Even when these time consuming analyses are performed by experienced cytogeneticists, it is difficult to pinpoint chromosomal breakpoints and segments involved in duplication, deletion, or translocation [l].We are attempting to develop procedures and DNA probe sets for visualization of specific numerical and structural aberrations in tumor cells. The detection of chromosome abnormalities in interphase nuclei, an approach termed "interphase cytogenetics"[3], appears to be a useful addition to the classical metaphase analysis. The examination of numerical aberrations in interphase nuclei can yield ' TO whom reprint requests should be addressed 199
A biotinylated Po glycoprotein cDNA was hybridized in situ to aldehyde-fixed vibratome sections and to aldehyde-fixed thin sections of Lowicryl-embedded trigeminal ganglia of 15 day old rats. Alkaline phosphatase and peroxidase detectors were used for light microscopic (LM) studies and peroxidase or colloidal gold were employed for electron microscopic (EM) detection. In both LM and EM sections, probe was found in cytoplasmic areas of myelin-forming Schwann cells that were enriched in granular endoplasmic reticulum, demonstrating that these regions contain Po mRNA. Interestingly, Po mRNA tended to cluster in regions close to the developing myelin sheath. Relatively simple methods are here described for EM detection of mRNA with reasonable tissue preservation and high resolution. These methods may be useful for developmental and disease-related studies of specific mRNAs in mammalian tissues.
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