Scalable Acid-Aided Lysis of Skin Samples Improves Proteome Coverage

Liquid chromatography-mass spectrometry is an evolving tool for comprehensive proteomic analyses across tissues. Despite the widespread use of liquid chromatography-mass spectrometry in dermatology, full-thickness human skin remains challenging to analyze. The skin extracellular matrix presents 2 major obstacles: the extensive crosslinking complicates protein extraction, and the high abundance of extracellular matrix proteins can mask lower-abundance proteins, reducing identification numbers. These limitations hinder progress in skin proteomics research. To address these challenges, we adapted the trifluoroacetic acid–based SPEED (Sample Preparation by Easy Extraction and Digestion) method for skin samples. Trifluoroacetic acid does not disrupt most crosslinks, allowing for the removal of abundant crosslinked extracellular matrix proteins. This enhanced proteome coverage, increasing the number of identified protein groups to over 6200 in healthy human skin. The improved sensitivity enabled the use of minimally invasive 2-mm punch biopsies, potentially facilitating greater patient enrolment than commonly used larger punch biopsies. We anticipate that these advancements in sample preparation will pave the way for analysis tools such as machine learning in skin proteomics, which require large datasets with high identification numbers. Furthermore, we extend this approach to tape strip proteomics and identify up to 2300 proteins in healthy skin, providing a cost-effective, scalable, and sensitive alternative to current workflows.