An Integreated Method for Mass Spectrometry-based proteomics analysis of Single Cell by using online sample preparation on a monolithic column

Cell heterogeneity exists in different cell types and different subcelluar populations. The studies in cell heterogeneity provide deep understanding in cancer therapy and drug responses. However, due to the low amount of proteins from single cell, single-cell characterization of metabolites, proteins, and post-translational modifications remains challenging. Thus, new methods for proteomics analysis of single cell are highly demanded. In this work, a monolithic column was prepared. The surface inside the capillary was modified with functional monomer for reverse phrase chromatography-based enrichment of proteins and subsequent online digestion. Then, cell contents were directly analyzed on electrospray ionization mass spectromety. This intergrated method was evaluated by internal label. iRT standard peptides with known mass were injected into the monolithic column as an intracellular primary peptides. The loss rate of the label peptide equals to the loss rate of proteins from single cell. The loss rate of integrated method was 70%. 790 proteins were identified from one Hela cell. According to the lower loss and shorter time of this method, it provides new tools for spatial detection of the human tissue.