Formaldehyde fixation helps preserve the proteome state during single-cell proteomics sample processing and analysis

Mass spectrometry-based single-cell proteomics (SCP) is gaining momentum but remains limited to a few laboratories due to the high costs and specialized expertise required. The ability to send samples to specialized core facilities would benefit non-specialist labs and popularize SCP for biological applications. However, no methods have been tested in SCP to "freeze" the proteome state while maintaining cell integrity for transfer between labs or prolonged sorting using fluorescence-activated cell sorting (FACS). This study evaluates whether short-term formaldehyde (FA) fixation can maintain cell states for shipping and on-site sorting, processing, and mass spectrometry analysis. We demonstrate that short-term FA fixation does not significantly affect protein recovery, even without heating and strong detergents, and maintains analytical depth compared to classical workflows. Fixation also preserves drug-induced specific perturbations of protein abundance during cell sorting and sample preparation for SCP analysis. Our findings suggest that formaldehyde FA fixation can facilitate SCP by enabling sample shipping and prolonged sorting, potentially democratizing access to SCP technology and expanding its application in biological research, thereby accelerating discoveries in cell biology and personalized medicine.