Protocol for low-input proteomic analysis of in situ fixed adult murine muscle stem cells

Abstract

Summary: Studying skeletal muscle stem cells (MuSCs) quiescence is challenging as they quickly activate within hours of isolation from muscle. Here, we present a protocol to disassociate and characterize fixed peptides from quiescent MuSCs using trapped ion-mobility time-of-flight mass spectrometry (MS). We describe steps for mouse perfusion, fluorescence-activated cell sorting preparation and sorting, protein extraction, digestion, and liquid chromatography MS analysis. This protocol can be applied to other less-abundant somatic stem cell types using mouse lines with a reporter.For complete details on the use and execution of this protocol, please refer to Zeng et al. (2022, 2023).1,2 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Keywords

• Mass Spectrometry
• Proteomics
• Stem Cells