Max Planck Institute for Brain Research, Max von Laue Strasse 4, 60438 Frankfurt, Germany.
Max Planck Institute of Biophysics, Max von Laue Strasse 3, 60438 Frankfurt, Germany.
STAR Protoc. 2021 Dec 22;3(1):101063. doi: 10.1016/j.xpro.2021.101063. eCollection 2022 Mar 18.
Cellular processes require tight and coordinated control of protein abundance, localization, and activity. One of the core mechanisms to achieve specific regulation of proteins is protein phosphorylation. Here we present a workflow to monitor protein abundance and phosphorylation in primary cultured neurons using liquid chromatography-coupled mass spectrometry. Our protocol provides a detailed guide on all steps for detection and label-free-quantification of phosphorylated and unmodified proteins of primary cortical neurons, including primary cell culture, phosphoproteomic sample preparation and data-processing, and evaluation. For complete details on the use and execution of this protocol, please refer to Desch et al. (2021).
细胞过程需要严格协调控制蛋白质的丰度、定位和活性。实现蛋白质特异性调节的核心机制之一是蛋白质磷酸化。在这里，我们介绍了一种使用液相色谱-串联质谱法监测原代培养神经元中蛋白质丰度和磷酸化的工作流程。本方案提供了关于检测和非标记定量原代皮质神经元中磷酸化和未修饰蛋白质的所有步骤的详细指南，包括原代细胞培养、磷酸蛋白质组样品制备和数据处理以及评估。有关该方案使用和执行的完整详细信息，请参阅 Desch 等人。(2021)。
