Images showing foci of γH2AX, 53BP1 and MRE11 DSB-marking proteins in nuclei of peripheral blood lymphocytes of minipigs.

(A–C) Images of minipig PBL immunostained for γH2AX (green), 53BP1 (red) and nuclei (DNA, blue; DAPI). (A) Control cells without IR exposure and DNA damage foci. (B) Minipig PBL 4 h post 49 Gy irradiation revealing fluorescent foci of γH2AX and 53BP1 marking DSBs. Foci usually contain variable amounts of both DSB markers leading to reddish – greenish colored foci due to color overlap (see insets in C,D). A cell with pan-γH2AX staining shows heavy overall green labeling (arrow). (C) PBL nuclei obtained 4 h post IR. A cell with 8 foci is seen in the image center (arrow). Inset: fluorescence intensity profiles across 5 foci in 3 PBL nuclei revealing colocalization of green γH2AX and red 53BP1 fluorescence intensity peaks. (D) PBLs of a 4 h sample showing colocalization of the DNA repair protein MRE11 (red) and γH2AX (green) at DNA damage foci, those appear whitish due to color overlap. The inset shows fluorescence intensity profiles across 4 foci in 2 PBL nuclei revealing colocalization of green γH2AX and red MRE11 fluorescence intensity peaks. Magnification 630× in the original micrographs.