Isolation and Flow Cytometric Analysis of Immune Cells from the Ischemic Mouse Brain

摘要: Ischemic stroke initiates a robust inflammatory response that starts in the intravascular compartment and involves rapid activation of brain resident cells. A key mechanism this is migration circulating immune cells to ischemic facilitated by chemokine release increased endothelial adhesion molecule expression. Brain-invading leukocytes are well-known contributing early-stage secondary injury, but their significance for termination inflammation later repair has only recently been noticed. Here, simple protocol efficient isolation from mouse provided. After transcardial perfusion, hemispheres dissected mechanically dissociated. Enzymatic digestion with Liberase is followed density gradient (such as Percoll) centrifugation remove myelin cell debris. One major advantage single-layer procedure which does not require time-consuming preparation gradients can be reliably performed. The approach yields highly reproducible counts per hemisphere allows measuring several flow cytometry panels one biological replicate. Phenotypic characterization quantification brain-invading after experimental may contribute better understanding multifaceted roles injury repair.