Fluorescent staining was performed to examine caspase‐3 activity and microglia activation after cerebral I/R as described previously.30 (link) Briefly, brains from each group were harvested and immersion‐fixed in 4% buffered parafomaldehyde, embedded in paraffin, cut at 7 μm, and stained with an specific anti‐cleaved caspase‐3 antibody (Cell Signaling Technology, Inc) or anti‐Iba‐1 antibody (Santa Cruz Biotechnology, Inc) as described previously.30 (link) After washing, the tissue sections were incubated with FITC‐conjugated anti‐rabbit (GeneTex) for 1 hour at 25°C and covered with fluorescence mounting medium (Life Technologies). The images were viewed on an EVOS‐fl digital inverted fluorescent microscopy (Advanced Microscopy Group). Fields of cortex were randomly examined using a defined rectangular field area for analysis of microglia activation. Total cells were counted in each field, and Iba‐1‐positive activated microglia cells are presented as the percentage of total cells counted.
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Caspase-3 and Microglia Activation in Cerebral I/R
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Anti antibody
Brains
Caspase 3
Cells
Cortex
Embedded paraffin
Fitc
Fluorescence
Immersion
Microglia
Microscopy
Rabbit
Tissue
Corresponding Organization :
Other organizations : East Tennessee State University, Nanjing Medical University, Jiangsu Province Hospital
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Variable analysis
independent variables
- Cerebral I/R
- Caspase-3 activity
- Microglia activation
- Sections were cut at 7 μm
- Staining was performed with specific anti-cleaved caspase-3 antibody and anti-Iba-1 antibody
- Tissue sections were incubated with FITC-conjugated anti-rabbit for 1 hour at 25°C
- Tissue sections were covered with fluorescence mounting medium
- Positive control: Not explicitly mentioned
- Negative control: Not explicitly mentioned
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