Figure 3.

Presence of intracellular and extracellular Hb. The presence of RBCs and Hb following IVH was characterized within the brain utilizing the inherent peroxidase activity of Hb in combination with SEM. Rabbit pups with IVH or sham controls were euthanized at 72 h of age followed by saline and freshly prepared 4% PFA perfusion. Afterwards the brains were dissected out from the skulls and immersed in 4% PFA. Brains were prepared and sections at the levels of rostral forebrain (Level 1) were stained for peroxidase activity of Hb or prepared for high magnification SEM as described in the Materials and Methods section. Bright-field microscope analyses were performed on a wide-field Olympus microscope (IX73) and slide scanning were performed on a Hamamatsu NanoZoomer 2.0-HT Digital slide scanner: C10730. Scanning was performed with a 40 × magnification lens. Images used for illustrations, from regions of interest (ROI), were grabbed with the viewer software NDP.view2 Viewing software. Scale bar of slide scan image indicate 1.0 mm and of ROI images indicate 100 μm. SEM specimens were examined in a Philips/FEI XL 30 FESEM scanning electron microscope using an Everhart-Tornley secondary electron detector. Image processing was done with the Scandium software for simple image acquiring and auto-storage into the Scandium database. Scale bar of SEM images indicate 10 μm.