Modifications to the plasma membrane proteome reflect neuronal physiological states. Here, we present a protocol to profile surface proteins from mouse cortical neuron cultures using in situ biotinylation. The workflow includes neuron culture, surface labeling, enrichment of biotinylated proteins, and on-beads digestion followed by mass spectrometry, enabling quantitative and comparative analysis of membrane-associated proteins across diverse physiological conditions. For complete details on the use and execution of this protocol, please refer to Shi et al. 1 • Instructions for labeling membrane proteins in situ in live neurons • Procedures for purifying membrane and membrane-associated proteins • Steps for identifying membrane proteins that report local membrane events Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. Modifications to the plasma membrane proteome reflect neuronal physiological states. Here, we present a protocol to profile surface proteins from mouse cortical neuron cultures using in situ biotinylation. The workflow includes neuron culture, surface labeling, enrichment of biotinylated proteins, and on-bead digestion followed by mass spectrometry, enabling quantitative and comparative analysis of membrane-associated proteins across diverse physiological conditions.
Shi et al. (Sun,) studied this question.