Protocol for the isolation of brain microvessels and visualization of RNA fluorescence in mice and humans

Olivia M Osborne; Oandy Naranjo; Silvia Torices; Sarah Schmidlin; Destiny Tiburcio; Minseon Park; Michal Toborek

doi:10.1016/j.xpro.2024.103530

Protocol for the isolation of brain microvessels and visualization of RNA fluorescence in mice and humans

STAR Protoc. 2025 Jan 7;6(1):103530. doi: 10.1016/j.xpro.2024.103530. Online ahead of print.

Authors

Olivia M Osborne¹, Oandy Naranjo², Silvia Torices³, Sarah Schmidlin³, Destiny Tiburcio³, Minseon Park³, Michal Toborek⁴

Affiliations

¹ University of Miami Miller School of Medicine, Department of Biochemistry and Molecular Biology, Miami, FL, USA. Electronic address: [email protected].
² University of Miami Miller School of Medicine, Department of Biochemistry and Molecular Biology, Miami, FL, USA. Electronic address: [email protected].
³ University of Miami Miller School of Medicine, Department of Biochemistry and Molecular Biology, Miami, FL, USA.
⁴ University of Miami Miller School of Medicine, Department of Biochemistry and Molecular Biology, Miami, FL, USA; Institute of Physiotherapy and Health Sciences, the Blood-Brain Barrier Research Center, the Jerzy Kukuczka Academy of Physical Education, Katowice, Poland. Electronic address: [email protected].

PMID: 39772387
DOI: 10.1016/j.xpro.2024.103530

Abstract

Here, we present a protocol for isolating microvessels from fresh or snap-frozen brain tissue from mice and humans, followed by visualization of RNA utilizing RNAscope hybridization for quantification of mRNA. We describe the steps for sample preparation and isolation, fixation, and hybridization. This protocol was specifically designed to integrate with RNAscope in situ hybridization. Although the protocol was developed in a mouse model, it can be optimized for use in other organisms, including human brain samples.

Keywords: Cell isolation; Classification Description; Gene Expression; Microscopy.