title: In-tip nanoreactors for cancer cells proteome profiling.
abstract: Mass spectrometry (MS)-based proteome profiling is essential for molecular diagnostics in modern biomedical study. To date, sample preparation including protein extraction and proteolysis is still very challenging and lack of efficiency. Recently tips-based sample preparation protocols exhibit strong potentials to achieve the goal of "a proteome in an hour". However, in-tip proteolysis is still rarely reported and far from ideal for dealing with complex bio-samples. In this work, nanoreactors encapsulated micropipette tips were demonstrated as high performance devices for fast (∼minutes) and multiplexing proteolysis to assist the profiling of cancer cells proteome. Nanoporous silica materials with controlled pore size and surface chemistry were prepared as nanoreactors and 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 proteomic analysis. 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 proteolysis based bottom-up proteomic strategy. With the enhanced performance, our work built a novel, efficient and miniaturized platform for facile proteomic sample preparation, which is promising for advanced biomarkers discovery in biomedical study.
authors.affiliation: Department of Chemistry, Institute of Biomedical Sciences, State Key Lab of Molecular Engineering of Polymers, Fudan University, Shanghai, 200433, China.
authors.forename: Ling
authors.initials: L
authors.lastname: Yan
authors.affiliation: Department of Chemistry, Institute of Biomedical Sciences, State Key Lab of Molecular Engineering of Polymers, Fudan University, Shanghai, 200433, China.
authors.forename: Liang
authors.initials: L
authors.lastname: Qiao
authors.affiliation: Department of Chemistry, Institute of Biomedical Sciences, State Key Lab of Molecular Engineering of Polymers, Fudan University, Shanghai, 200433, China.
authors.forename: Ji
authors.initials: J
authors.lastname: Ji
authors.affiliation: Department of Chemistry, Institute of Biomedical Sciences, State Key Lab of Molecular Engineering of Polymers, Fudan University, Shanghai, 200433, China.
authors.forename: Yixin
authors.initials: Y
authors.lastname: Li
authors.affiliation: Department of Chemistry, Institute of Biomedical Sciences, State Key Lab of Molecular Engineering of Polymers, Fudan University, Shanghai, 200433, China.
authors.forename: Xuefei
authors.initials: X
authors.lastname: Yin
authors.affiliation: Department of Chemistry, Institute of Biomedical Sciences, State Key Lab of Molecular Engineering of Polymers, Fudan University, Shanghai, 200433, China.
authors.forename: Ling
authors.initials: L
authors.lastname: Lin
authors.affiliation: Department of Chemistry, Institute of Biomedical Sciences, State Key Lab of Molecular Engineering of Polymers, Fudan University, Shanghai, 200433, China.
authors.forename: Xiaohui
authors.initials: X
authors.lastname: Liu
authors.affiliation: Department of Chemistry, Institute of Biomedical Sciences, State Key Lab of Molecular Engineering of Polymers, Fudan University, Shanghai, 200433, China.
authors.forename: Jun
authors.initials: J
authors.lastname: Yao
authors.affiliation: Department of Chemistry, Institute of Biomedical Sciences, State Key Lab of Molecular Engineering of Polymers, Fudan University, Shanghai, 200433, China.
authors.forename: Yi
authors.initials: Y
authors.lastname: Wang
authors.affiliation: School of Biomedical Engineering, Med-X Research Institute, Shanghai Jiao Tong University, Shanghai, 200030, China.
authors.forename: Bin
authors.initials: B
authors.lastname: Liu
authors.affiliation: School of Biomedical Engineering, Med-X Research Institute, Shanghai Jiao Tong University, Shanghai, 200030, China.
authors.forename: Kun
authors.initials: K
authors.lastname: Qian
authors.affiliation: Department of Chemistry, Institute of Biomedical Sciences, State Key Lab of Molecular Engineering of Polymers, Fudan University, Shanghai, 200433, China.
authors.forename: Baohong
authors.initials: B
authors.lastname: Liu
authors.affiliation: Department of Chemistry, Institute of Biomedical Sciences, State Key Lab of Molecular Engineering of Polymers, Fudan University, Shanghai, 200433, China. Electronic address: pyyang@fudan.edu.cn.
authors.forename: Pengyuan
authors.initials: P
authors.lastname: Yang
mesh_terms: D045744:Cell Line, Tumor; D006801:Humans; D013058:Mass Spectrometry; D009369:Neoplasms; D059748:Proteolysis; D020543:Proteome / Q000378:metabolism*; D040901:Proteomics
chemical_list: D020543:Proteome
keywords: Cancer cells; Mass spectrometry; Micropipette tips; Nanoporous materials; Profiling; Proteolysis