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3d Confocal Microscopy and Image Analysis for Measurement of Genetic Instability

Published online by Cambridge University Press:  02 July 2020

C. Ortiz de Solorzano ,
K. Chin ,
D. Knowles ,
A. Jones ,
E. Garcia ,
J.W. Gray  and
S.J. Lockett
C. Ortiz de Solorzano
Affiliation:
Genome Sciences Department, Life Sciences Division. Lawrence Berkeley National Laboratory. University of California, BerkeleyCA94720
K. Chin
Affiliation:
Cancer Center, University of California, San FranciscoCA94115
D. Knowles
Affiliation:
Genome Sciences Department, Life Sciences Division. Lawrence Berkeley National Laboratory. University of California, BerkeleyCA94720
A. Jones
Affiliation:
Genome Sciences Department, Life Sciences Division. Lawrence Berkeley National Laboratory. University of California, BerkeleyCA94720
E. Garcia
Affiliation:
Genome Sciences Department, Life Sciences Division. Lawrence Berkeley National Laboratory. University of California, BerkeleyCA94720
J.W. Gray
Affiliation:
Cancer Center, University of California, San FranciscoCA94115
S.J. Lockett
Affiliation:
Genome Sciences Department, Life Sciences Division. Lawrence Berkeley National Laboratory. University of California, BerkeleyCA94720
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Extract

Solid tumors frequently contain cells that are heterogeneous in the copy number of DNA loci. This fact implies the existence of genetic instability, which may be associated with disease aggressiveness. Accurate measurement of this phenomenon requires analysis of intact nuclei within their natural tissue context. We perform these measurements by preparing >30μm thick tissue sections, labeling them with fluorescent labels for total DNA and for specific DNA loci using fluorescence in situ hybridization (FISH) which retain the transparency of the tissue and acquiring 3D images of the tissue using confocal microscopy (figure 1). In this study, we combined automated 3D image analysis (IA) algorithms for segmenting individual nuclei based on the total DNA stain1 and for segmenting the punctuate FISH signals of DNA loci. This enables us to efficiently enumerate the copy number of specific DNA loci in individual cells and as a function of the cell's location in the tissue.

Type
Structural Approaches to the Study of Cell Cell Interactions In Three Dimensions
Information
Microscopy and Microanalysis , Volume 5 , Issue S2: Proceedings: Microscopy & Microanalysis '99, Microscopy Society of America 57th Annual Meeting, Microbeam Analysis Society 33rd Annual Meeting, Portland, Oregon August 1-5, 1999 , August 1999 , pp. 1022 - 1023
Copyright
Copyright © Microscopy Society of America

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References

1.Ortiz de Solorzano, C. et al. Journal of Microscopy 193(1999)212.CrossRefGoogle Scholar
2.Ortiz de Solorzano, C. et al. Cytometry 31(1998)93.3.0.CO;2-J>CrossRefGoogle Scholar
3.Lockett, S.J. et ai, Proc. Ann. MSA Meeting 55(1997)1121.Google Scholar
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