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September 5, 2025Open Access

Nuclei Isolation Protocol for Single-Nucleus RNA Sequencing of Human Stem Cell-Derived Grafts

ESEdoardo SozziLund University PSPetter StormLund University AFAlessandro FiorenzanoLund University

Puntos clave

This protocol enhances single-nucleus RNA sequencing for assessing human grafts in the rat brain.
Key techniques include vibratome sectioning and fluorescence-activated sorting to extract nuclei effectively.
It facilitates in-depth analysis of neuronal identity and graft integration in a neurodegeneration context.
This method supports insights into complex interactions within the host brain tissue and transplanted cells.

Resumen

Single-nucleus RNA sequencing enables high-resolution transcriptomic profiling of brain tissue, facilitating detailed analysis of cell identity in models of neurodegeneration and repair. Here, we describe a protocol for isolating nuclei from long-term human stem cell-derived grafts in the rat brain, incorporating vibratome sectioning, graft dissection, nuclear extraction, and fluorescence-activated sorting. This workflow supports analysis of human neurons embedded within host tissue or sensitive to dissociation, offering a powerful approach to assess graft composition, integration, and neuronal identity in living brain.

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