In-tip nanoreactors for cancer cells proteome profiling

Abstract Mass spectrometry (MS)-based proteomeprofiling is essential for molecular diagnostics in modern biomedical study. To date, sample preparation including protein extraction and proteolysisis still very challenging and lack of efficiency. Recently tips-based sample preparation protocols exhibit strong potentials to achieve the goal of “a proteomein an hour”. However, in-tip proteolysisis still rarely reported and far from ideal for dealing with complex bio-samples. In this work, nanoreactorsencapsulated micropipette tips were demonstrated as high performance devices for fast (∼minutes) and multiplexing proteolysisto assist the profiling of cancer cells proteome. Nanoporous silica materials with controlled pore size and surface chemistry were prepared as nanoreactorsand encapsulated in micropipette tips for efficient in situ proteolysis. The as-constructed device showed desirable sensitivity (LOD of 0.204 ± 0.008 ng/μL and LOQ of 0.937 ± 0.055 ng/μL), selectivity, stability (two months under −20 °C), reusability (at least 10 times), and little memory effect in MS based bottom-up proteomicanalysis. It was used for comprehensive protein mapping from cancer cell lines. The number of identified proteins was increased by 18%, 22%, 52%, and 52% dealing with HepG2, F56, MCF7, and HCCLM3 cancer cells, compared to traditional in-solution proteolysisbased bottom-up proteomicstrategy. With the enhanced performance, our work built a novel, efficient and miniaturized platform for facile proteomicsample preparation, which is promising for advanced biomarkers discoveryin biomedical study.