Gamma-H2AX foci counting: image processing and control software for high-content screening

P. R. Barber; R. J. Locke; G. P. Pierce; K. Rothkamm; B. Vojnovic

doi:10.1117/12.705217

19 February 2007 Gamma-H2AX foci counting: image processing and control software for high-content screening

P. R. Barber, R. J. Locke, G. P. Pierce, K. Rothkamm, B. Vojnovic

Proceedings Volume 6441, Imaging, Manipulation, and Analysis of Biomolecules, Cells, and Tissues V; 64411M (2007) https://doi.org/10.1117/12.705217
Event: SPIE BiOS, 2007, San Jose, California, United States

Abstract

Phosphorylation of the chromatin protein H2AX (forming γH2AX) is implicated in the repair of DNA double strand breaks (DSB's); a large number of H2AX molecules become phosphorylated at the sites of DSB's. Fluorescent staining of the cell nuclei for γH2AX, via an antibody, visualises the formation of these foci, allowing the quantification of DNA DSB's and forming the basis for a sensitive biological dosimeter of ionising radiation. We describe an automated fluorescence microscopy system, including automated image processing, to count γH2AX foci. The image processing is performed by a Hough transform based algorithm, CHARM, which has wide applicability for the detection and analysis of cells and cell colonies. This algorithm and its applications for cell nucleus and foci detection will be described. The system also relies heavily on robust control software, written using multi-threaded cbased modules in LabWindows/CVI that adapt to the timing requirements of a particular experiment for optimised slide/plate scanning and mosaicing, making use of modern multi-core processors. The system forms the basis of a general purpose high-content screening platform with wide ranging applications in live and fixed cell imaging and tissue micro arrays, that in future, can incorporate spectrally and time-resolved information.

Citation Download Citation

P. R. Barber, R. J. Locke, G. P. Pierce, K. Rothkamm, and B. Vojnovic "Gamma-H2AX foci counting: image processing and control software for high-content screening", Proc. SPIE 6441, Imaging, Manipulation, and Analysis of Biomolecules, Cells, and Tissues V, 64411M (19 February 2007); https://doi.org/10.1117/12.705217

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