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Extraction and Tissue Clearing Preparation of Mouse Brain-Spinal Cord Samples
Published on: April 11, 2025
Brain Tissue Sample Stabilization and Extraction Strategies for Neuropeptidomics
Elva Fridjonsdottir1, Anna Nilsson1, Henrik Wadensten1
1Biomolecular Imaging and Proteomics, Department of Pharmaceutical Biosciences, Uppsala University, Uppsala, Sweden.
Abstract:
Neuropeptides are bioactive peptides that are synthesized and secreted by neurons in signaling pathways in the brain. Peptides and proteins are extremely vulnerable to proteolytic cleavage when their biological surrounding changes. This makes neuropeptidomics challenging due to the rapid alterations that occur to the peptidome after harvesting of brain tissue samples. For a successful neuropeptidomic study the biological tissue sample analyzed should resemble the premortem state as much as possible. Heat stabilization has been proven to inhibit postmortem degradation by denaturing proteolytic enzymes, hence increasing identification rates of neuropeptides. Here, we describe a stabilization protocol of a frozen tissue specimen that increases the number of intact mature neuropeptides identified and minimizes interference of degradation products from abundant proteins. Additionally, we present an extraction protocol that aims to extract a wide range of hydrophilic and hydrophobic neuropeptides by using both an aqueous and an organic extraction medium.
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