Immuno-laser capture microdissection of perfusion-fixed brain tissue coupled to RNA-seq

Normal mouse brain was fixed by cardiac perfusion with 4% paraformaldehyde in PBS, then post-fixed by immersion in the same solution for 2hr at RT. Brain was embedded in cryomatrix and frozen sections were cut at 7 um on a cryostat. Sections were immunostained with anti-CD31 antibody and anti-GFAP antibody to detect microvascular endothelial cells and surrounding astrocytes, respectively. Immediately after immunostaining, sections were subject to laser capture microdissection (LCM), using ultraviolet laser to ablate unwanted astrocytes and infrared laser to lift target endothelial tissue. Total RNA was isolated from LCM-retrieved endothelial tissue and processed for RNA-seq. RNA-seq profiling of Immuno-laser capture microdissection of perfusion-fixed brain tissue