Mak115

Total protein content of samples (20 µL) was assayed according to Bradford (34 (link)). Drabkin's reagent was used to measure hemoglobin concentration in blood hemolysates using the assay kit MAK 115 purchased from Sigma-Aldrich (St. Louis, MO, USA). Ferricyanide oxidizes the iron in hemoglobin thereby changing it to methemoglobin which binds with potassium cyanide to form cyanmethemoglobin that absorbs light at 540 nm.
Statistical analysis was performed using the computer based package of statistical product and service solution (SPSS version 9.0). Sample analysis was run in duplicate for all investigated parameters and results presented as means±SD. Values of the activities and concentrations of individual parameters were compared between different groups of the study subjects using one-way analysis of variance (ANOVA) followed by the post hoc Tukey-HSD test for multiple comparisons. Values of P<0.05 were considered to be statistically significant.

The mice were euthanized by deep anesthesia, and coronal brain sections (1 mm thick) were cut with a mouse brain slice mold (40–75, RWD). The sections were digitized and analyzed with ImageJ (Fiji, 2.1.0/1.53c, NIH Image, USA). We photographed all brain sections of each mouse for quantification. The total volume of hematoma was calculated and used for statistical analysis.
The hemoglobin content was measured with a hemoglobin assay kit (MAK115, Sigma-Aldrich) according to the manufacturer’s instructions.⁷³ (link) Briefly, 4-mm thick brain tissue from the ipsilateral striatum, including the hematoma core and perihematomal region, was collected and homogenized for 3 min by sonication in 300 μL of ice-cold phosphate-buffered saline (PBS). After the homogenate was centrifuged, 50 μL of the supernatant was mixed with 200 μL hemoglobin detection reagent. The mixture was incubated for 5minat room temperature, and the absorbance was measured at 400 nm. The optical density of each sample was plotted on a standard curve.