Comparison of two methods for measuring gamma-H2AX nuclear fluorescence as a marker of DNA damage in cultured human cells: applications for microbeam radiation therapy
2013 (English)In: Journal of Instrumentation, ISSN 1748-0221, Vol. 8, C06008- p.Article in journal (Refereed) Published
Microbeam radiation therapy (MRT) delivers single fractions of very high doses of synchrotron x-rays using arrays of microbeams. In animal experiments, MRT has achieved higher tumour control and less normal tissue toxicity compared to single-fraction broad beam irradiations of much lower dose. The mechanism behind the normal tissue sparing of MRT has yet to be fully explained. An accurate method for evaluating DNA damage, such as the gamma-H2AX immunofluorescence assay, will be important for understanding the role of cellular communication in the radiobiological response of normal and cancerous cell types to MRT. We compare two methods of quantifying gamma-H2AX nuclear fluorescence for uniformly irradiated cell cultures: manual counting of gamma-H2AX foci by eye, and an automated, MATLAB-based fluorescence intensity measurement. We also demonstrate the automated analysis of cell cultures irradiated with an array of microbeams. In addition to offering a relatively high dynamic range of gamma-H2AX signal versus irradiation dose (>10 Gy), our automated method provides speed, robustness, and objectivity when examining a series of images. Our in-house analysis facilitates the automated extraction of the spatial distribution of the gamma-H2AX intensity with respect to the microbeam array - for example, the intensities in the peak (high dose area) and valley (area between two microbeams) regions. The automated analysis is particularly beneficial when processing a large number of samples, as is needed to systematically study the relationship between the numerous dosimetric and geometric parameters involved with MRT (e.g., microbeam width, microbeam spacing, microbeam array dimensions, peak dose, valley dose, and geometric arrangement of multiple arrays) and the resulting DNA damage.
Place, publisher, year, edition, pages
2013. Vol. 8, C06008- p.
Radiotherapy concepts, Radiation damage evaluation methods, microbeam, H2AX foci
IdentifiersURN: urn:nbn:se:su:diva-92815DOI: 10.1088/1748-0221/8/06/C06008ISI: 000321627400008OAI: oai:DiVA.org:su-92815DiVA: diva2:642524