Stereological Analysis of Cell Counts

Tissue samples were prepared as described above and stereological counts were performed as previously described using a motorized Olympus BX51TRF microscope, Optronix cooled video camera and MicroBrightField StereoInvestigator software package (MBF Bioscience, Williston, VT, USA) (Tsenkina et al., 2015 (link)). Nonbiased ‘blinded’ cell number evaluation was performed by means of the optical fractionator method and the optical dissector probe. White and grey matter regions of interest in the ipsilateral hemisphere (depicted in Fig. 1) were manually traced with contours using 5× magnification. Subsequently, a grid of 200 × 200 µm² was placed over the selected area and immunopositive cells were counted using an optical fractionator at 63× (sampling box 50 × 50 µm²) at 2 hpi, 6 hpi, 24 hpi and/or 7 days post-injury (dpi) (Supplementary Fig. 1A and B).