in vivo TurboID

This protocol describes an astrocyte-specific in vivo proximity biotinylation (iBioID) approach using a Lamp1-TurboID fusion protein to investigate protein interactions within the endo-/lysosomal compartment of astrocytes. By leveraging astrocyte-specific AAV delivery in neonatal (P1) mice, this method enables selective biotinylation of proteins in a physiologically relevant context while preserving interactions with neighboring cells. Following biotin administration, labeled proteins are purified using streptavidin affinity capture and analyzed via quantitative LC-MS/MSto map astrocytic endolysosomal protein networks. This approach provides insights into subcellular proteomes and dynamic astrocyte signaling in vivo.