# Protocol for generating protein profiles and distance-based network analysis of murine tissue slices

## TL;DR

This paper presents a detailed protocol for spatial proteomics using mouse skeletal muscle tissue to generate protein profiles and analyze protein networks.

## Contribution

A new protocol for spatial proteomics using LC-MS/MS and a sliding window approach for missing data in tissue sections.

## Key findings

- The protocol enables spatial protein profiling along the longitudinal axis of skeletal muscle.
- A sliding window method effectively replaces missing data in protein profiles.
- The approach is applicable to other asymmetric tissues like the brain and heart.

## Abstract

We introduce a protocol for spatial proteomics using thin cryotome sections of mouse skeletal muscle tissue. We describe steps for preparing muscle sections and liquid chromatography-tandem mass spectrometry (LC-MS/MS) analyses to generate spatial protein profiles along the longitudinal skeletal muscle axis. We detail procedures for scanning longitudinal protein profiles and replacing missing data using a sliding window approach. This protocol has potential for applications in spatial proteomics to other tissues with asymmetric patterns such as the brain and heart tissue.

For complete details on the use and execution of this protocol, please refer to Schmidt et al.1

•Protocol for generating protein profiles based on LC-MS/MS data from tissue sections•Simple steps for sample preparation for LC-MS/MS analysis from tissue sections•Detailed instructions for data analysis to generate protein profiles•Precise guidelines for network generation of protein profiles

Protocol for generating protein profiles based on LC-MS/MS data from tissue sections

Simple steps for sample preparation for LC-MS/MS analysis from tissue sections

Detailed instructions for data analysis to generate protein profiles

Precise guidelines for network generation of protein profiles

Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

We introduce a protocol for spatial proteomics using thin cryotome sections of mouse skeletal muscle tissue. We describe steps for preparing muscle sections and liquid chromatography-tandem mass spectrometry (LC-MS/MS) analyses to generate spatial protein profiles along the longitudinal skeletal muscle axis. We detail procedures for scanning longitudinal protein profiles and replacing missing data using a sliding window approach. This protocol has potential for applications in spatial proteomics to other tissues with asymmetric patterns such as the brain and heart tissue.