Analysis of target mRNAs in the fixed-frozen human brain using a modified BaseScope-ISH Assay protocol

Abstract

Summary: We describe a modified BaseScope™ Assay protocol (ACDBio) for RNA in situ hybridization on fixed-frozen human brain tissue. The original protocol caused tissue detachment due to harsh tissue pre-treatment. We therefore optimized it to improve tissue stability while providing high stain quality in fragile post-mortem tissue from aged donors with advanced neurodegeneration. The main changes include two additional fixation steps and modifications to the pre-treatment protocol. We also describe tissue imaging and stain quantification using the open-source QuPath software.For complete details on the use and execution of this protocol, please refer to Hornsby et al. (2020). : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Keywords

• Health Sciences
• Microscopy
• In Situ Hybridization
• Neuroscience