A highly efficient method for extracting peptides from a single mouse hypothalamus

Abstract

Living organisms contain a variety of endogenous peptides that function as significant regulators of many biological processes. Endogenous peptides are typically analyzed using liquid chromatography–mass spectrometry (LC-MS). However, due to the low efficiency of peptide extraction and low abundance of peptides in a single animal, LC-MS–based peptidomics studies have not facilitated an understanding of the individual differences and tissue specificity of peptide abundance. In this study, we developed a peptide extraction method followed by nano-flow LC-MS/MS analysis. This method enabled highly efficient and reproducible peptide extraction from sub-milligram quantities of hypothalamus dissected from a single animal. Diverse bioactive and authentic peptides were detected from a sample volume equivalent to 135 μg of hypothalamus. This method may be useful for elucidating individual differences and tissue specificity, as well as for facilitating the discovery of novel bioactive peptides and biomarkers and developing peptide therapeutics.