Department of Neurology, Yale University School of Medicine, New Haven, CT 06520, USA.
Interdepartmental Neuroscience Program, Yale University, New Haven, CT 06520, USA.
STAR Protoc. 2021 Nov 24;2(4):100941. doi: 10.1016/j.xpro.2021.100941. eCollection 2021 Dec 17.
This protocol provides an improved pipeline for dissociating intact projection neurons from adult mouse cortex for applications including droplet and plate-based single-cell RNA sequencing, qPCR, immunocytochemistry, and long-term cell culture. This protocol provides a robust and reproducible dissociation pipeline that uses exclusively off-the-shelf reagents, not requiring the use of expensive dissociation kits. The unique incubation steps, in combination with the FACS gating strategy, results in unparalleled enrichment for intact cortical neurons from the adult brain. For complete details on the use and execution of this protocol, please refer to Golan et al. (2021).
本方案提供了一种改良的从成年小鼠皮层分离完整投射神经元的方案，适用于包括液滴和板基单细胞 RNA 测序、qPCR、免疫细胞化学和长期细胞培养等应用。本方案提供了一个强大且可重复的分离方案，仅使用现成的试剂，不需要使用昂贵的解离试剂盒。独特的孵育步骤，结合 FACS 门控策略，可从成年大脑中获得无与伦比的完整皮质神经元富集。有关本方案使用和实施的完整详细信息，请参阅 Golan 等人（2021 年）。
