After 6 h stroke in mice, ischemic and non-ischemic hemispheres of brains were snap frozen in liquid nitrogen and stored at −80 °C until further use for immunoblotting of fibrin(ogen) and MMP-9 [26 (link)-28 (link)]. Brain tissue (50 mg) was homogenized in lysis buffer (0.5 M Tris-HCl, pH 7.4, 1.5 M NaCl, 2.5% deoxycholic acid, 10% NP-40, 10 mM EDTA) containing 1.5 % SDS and 25 mM EACA (ε-aminocaproic acid) (for plasmin inhibition). Protein was estimated by bicinchoninic acid assay using homogenized tissue supernatant after centrifugation. Proteins (50 μg) were electrophoresed on 10% reducing SDS-PAGE gels and transferred to polyvinylidenedifluoride membrane. The membranes were blocked with 1% BSA in PBS containing 0.1% Tween 20. Fibrin(ogen) and MMP-9 were probed with rabbit anti-mouse fibrinogen antibody (MyBiosource) at 1:3000 dilution and goat anti-mouse MMP-9 (1:1000) antibodies respectively, overnight at 4 °C and, detected with anti-rabbit and anti-goat IRDye Li-COR secondary antibodies (LI-COR biosciences, NE). The membranes were scanned on LI-COR Odyssey scanner and pixel density was measured by NIH Image J software. Gelatinase zymography was performed as described [26 (link)].; additional details are available in the Supplemental Data.