Isolation of Capillaries from Small Amounts of Mouse Brain Tissue.

ABSTRACT

The integrity of the blood-brain barrier (BBB) is essential for the normal functioning of the central nervous system (CNS). Isolated brain capillaries are essential for analyzing changes in protein and gene expression at the BBB under physiological and pathological conditions. The standard methods for isolating brain capillaries require the use of at least one or more mouse brains in order to obtain sufficient quantity and purity of brain capillaries. Here, we describe an optimized protocol for isolating and purifying capillaries from tiny amounts of mouse cerebral cortex using manual homogenization, density gradient centrifugation, and filtration while preserving the structural integrity and functional activity of microvessel fragments. Western blotting showed that proteins expressed at the BBB were enriched in mouse brain capillaries isolated by the optimized method compared to cerebral cortex protein homogenates. This approach can be used for the analysis of a variety of rare mouse genetic models and can also help the investigators to understand regional differences in susceptibility to pathological phenomena such as ischemia and traumatic brain injury. This will allow the investigators to better understand the physiology and pathology of the BBB.