FociCounter: a freely available PC programme for quantitative and qualitative analysis of gamma-H2AX foci.

Gamma-H2AX foci are sensitive and specific indicator for the induction of DNA double-strand breaks (DSBs) and an immunocytochemical assay with antibodies recognizing gamma-H2AX has become the gold standard for the detection of this type of DNA lesion. Quantification of gamma-H2AX foci can be achieved by various methods such as Western blotting, flow cytometry, visual analysis and computational analysis with a fluorescence microscope. The best sensitivity is achieved by computer analysis. Since no freeware programme for the analysis of gamma-H2AX foci exists for a PC platform, the aim of our study was to develop a simple and user-friendly public-domain software. The algorithm applied in our programme allows determination of the number of foci in a single cell, a focus intensity per cell, as well as a cell intensity. Its graphical user interface is based on a GTK+ library and the whole application can be run under a variety of operating systems, including MS Windows and Linux. The programme called FociCounter is publicly available at http://focicounter.sourceforge.net. Application of the programme was tested by analysing gamma-H2AX foci in CHO and MO59K cells irradiated in vitro with X-rays and validated by comparing the results obtained with the outcome of automated image analysis and flow cytometry.

[1]  Michel Nussenzweig,et al.  H2AX: the histone guardian of the genome. , 2004, DNA repair.

[2]  Matthew A. Coleman,et al.  Candidate protein biodosimeters of human exposure to ionizing radiation , 2006, International journal of radiation biology.

[3]  E. Rogakou,et al.  Histone H2A variants H2AX and H2AZ. , 2002, Current opinion in genetics & development.

[4]  George Iliakis,et al.  Computational Methods for Analysis of Foci: Validation for Radiation-Induced γ-H2AX Foci in Human Cells , 2006, Radiation research.

[5]  P. Olive,et al.  Phosphorylation of histone H2AX as a measure of radiosensitivity. , 2004, International journal of radiation oncology, biology, physics.

[6]  Takeo Ohnishi,et al.  Does gammaH2AX foci formation depend on the presence of DNA double strand breaks? , 2005, Cancer letters.

[7]  J. Lieberman,et al.  A phosphatase complex that dephosphorylates γH2AX regulates DNA damage checkpoint recovery , 2006, Nature.

[8]  Takeo Ohnishi,et al.  Does γH2AX foci formation depend on the presence of DNA double strand breaks , 2005 .

[9]  A. Hauptner,et al.  Methods for quantitative evaluation of dynamics of repair proteins within irradiated cells , 2006 .

[10]  Borivoj Vojnovic,et al.  Gamma-H2AX foci counting: image processing and control software for high-content screening , 2007, SPIE BiOS.

[11]  Michael Uder,et al.  In vivo formation and repair of DNA double-strand breaks after computed tomography examinations. , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[12]  Vicky Goh,et al.  Leukocyte DNA damage after multi-detector row CT: a quantitative biomarker of low-level radiation exposure. , 2007, Radiology.

[13]  T. Lörch,et al.  New developments in automated cytogenetic imaging: unattended scoring of dicentric chromosomes, micronuclei, single cell gel electrophoresis, and fluorescence signals , 2004, Cytogenetic and Genome Research.

[14]  E. Rogakou,et al.  DNA Double-stranded Breaks Induce Histone H2AX Phosphorylation on Serine 139* , 1998, The Journal of Biological Chemistry.

[15]  Kai Rothkamm,et al.  Evidence for a lack of DNA double-strand break repair in human cells exposed to very low x-ray doses , 2003, Proceedings of the National Academy of Sciences of the United States of America.