C18-Functionalized Amine-Bridged Hybrid Monoliths for Mass Spectrometry-Friendly Peptide Separation and Highly...

For proteomic analysis based on mass spectrometry (MS), high-performance peptide separation under MS-friendly conditions is of importance. To this end, a novel kind of amine-bridged hybrid monolith was developed by the sol-gel reaction of bis[3-(trimethoxysilyl)propyl]amine and allyltrimethoxysilane, followed by "thiol-ene" click functionalization of C18 groups. With the secondary amino groups bridged in the framework, the nonspecific adsorption from silanol groups could be decreased, so that peptide peak tailing under MS-friendly conditions was reduced, and half peak width was narrowed. Furthermore, such materials were facilely in situ prepared in the very narrow bore capillary with low backpressure for proteomic analysis of limited amounts of samples. Finally, 16,692 unique peptides corresponding to 3698 protein groups could be averagely identified from 10 ng Hela cell digests in a single 65 min run, and 5257 peptides corresponding to 1062 protein groups could be averagely identified from 200 pg digests in a single 60 min run. Such high sensitivity could be attributed to the decreased nonspecific adsorption, the narrowed peak width, and the miniaturization of the column. It is shown that such monoliths are promising for highly sensitive proteomic analysis, including single-cell proteomics.