Objective: To test a CellProfiler pipeline for automated counting and characterization of gamma-H2AX foci in color images of human cultured cells.
Study design: A431 cells were irradiated and stained for gamma-H2AX foci detection. Sets of color images were analyzed visually, and findings were compared with those using FociCounter and CellProfiler software.
Results: The CellProfiler pipeline includes some proprieties not present in FociCounter, such as the automatic detection of nuclei, the detection of touching nuclei and the rejection of nuclei that touch the border of the image. The time required for manual operation is associated with the number of images analyzed visually or by FociCounter but not for the CellProfiler program. CellProfiler reduced manual operation time and is about 4 times faster than semiautomatic detection using FociCounter and 10 times faster than visual counting.
Conclusion: We conclude that CellProfiler and FociCounter are reliable tools for measuring gamma-H2AX foci. However, CellProfiler overcomes the limitations of the FociCounter program and is able to detect nuclei automatically, saving considerable manual operation.