Interactive algorithms for rapid enumeration of hybridization signals in individual whole-cell nuclei inside intact-tissue specimens

Stephen J. Lockett, Curtis T. Thompson, James C. Mullikin, Damir Sudar, R. Khavari, W. Hyun, Daniel Pinkel, Joe W. Gray

Research output: Chapter in Book/Report/Conference proceedingConference contribution

2 Scopus citations

Abstract

Fluorescence in situ hybridization (FISH) is useful for analyzing specific nucleic acid sequences in individual cells. Its application to tissue sections has been limited however because of the difficulties of performing the hybridization and analysis in sections that are thick enough to contain intact nuclei. Recent improvements in FISH permit hybridization with chromosome-specific, centromeric probes throughout 20 μm formalin fixed, paraffin- embedded sections, which do contain many intact nuclei. This paper describes software to facilitate analysis of these 3D hybridizations. We have developed two algorithms for analyzing 3D, confocal images of thick sections. One displays 2D, maximum-intensity, projection images through the original 3D image at different angles. When projections are viewed sequentially, the 3D image appears semi-transparent and rotates. The second algorithm allows interactive enumeration of FISH signals. Each signal is marked by the analyst. Then, for each pair of marked signals, a 2D slice image along the line connecting both marked signals and parallel to the z (depth) axis is displayed. From this slice, the analyst decides if the signals are in the same or different nuclei, or if the signals should be rejected because they are in a nucleus truncated by the upper or lower surface of the section. After consideration of all pairs of signals, the algorithm produces a map of the tissue section showing the numbers of signals in each of the intact nucleus. The algorithms enable analysis of small, premalignant and early malignant lesions and infiltrative lesions that cannot be analyzed by other molecular techniques and permit the direct correlation of FISH information with histology/cytology.

Original languageEnglish (US)
Title of host publicationProceedings of SPIE - The International Society for Optical Engineering
EditorsTony Wilson, Carol J. Cogswell
PublisherSociety of Photo-Optical Instrumentation Engineers
Pages43-49
Number of pages7
ISBN (Print)0819417599
StatePublished - Jan 1 1995
EventThree-Dimensional Microscopy: Image Acquisition and Processing II - San Jose, CA, USA
Duration: Feb 9 1995Feb 10 1995

Publication series

NameProceedings of SPIE - The International Society for Optical Engineering
Volume2412
ISSN (Print)0277-786X

Other

OtherThree-Dimensional Microscopy: Image Acquisition and Processing II
CitySan Jose, CA, USA
Period2/9/952/10/95

ASJC Scopus subject areas

  • Electronic, Optical and Magnetic Materials
  • Condensed Matter Physics
  • Computer Science Applications
  • Applied Mathematics
  • Electrical and Electronic Engineering

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  • Cite this

    Lockett, S. J., Thompson, C. T., Mullikin, J. C., Sudar, D., Khavari, R., Hyun, W., Pinkel, D., & Gray, J. W. (1995). Interactive algorithms for rapid enumeration of hybridization signals in individual whole-cell nuclei inside intact-tissue specimens. In T. Wilson, & C. J. Cogswell (Eds.), Proceedings of SPIE - The International Society for Optical Engineering (pp. 43-49). (Proceedings of SPIE - The International Society for Optical Engineering; Vol. 2412). Society of Photo-Optical Instrumentation Engineers.