The proteome reflects the physiology and pathology states of a patient therefore proteomics is a powerful tool for early diagnostics of diseases and monitoring of therapeutic responses. Mass spectrometry (MS) measures the mass-to-charge ratio of charged species and has become the enabling technology for proteomics. However, the majority of the current proteomics studies rely on bottom-up/shotgun approaches. In this case, mixtures of proteins are digested by one of the proteases (e.g., trypsin), separated by liquid chromatography (LC), and analyzed by electrospray mass spectrometry (ESI-MS). Despite tremendous successes, there remain two major limitations in bottom-up proteomics: first, it is difficult to identify all protein isoforms or proteoforms, including splicing, modifications, cleavages, etc.; second, the native state of proteins is always lost after digestion. There is currently a great push to implement top-down proteomics, i.e., identification and characterization of full-length proteins by LC-MS. Unfortunately, top-down proteomics proves to be much more challenging. There are several bottlenecks: first, lower MS sensitivity of protein relative to peptides; second, limitation on detection of high molecular weight proteins; third, inefficient identification of proteins by MS/MS fragmentation; and fourth, laborious multidimensional protein separation not suitable for small volumes of biological samples. The field is calling for transformative technologies. In response to PA-11-215, Newomics Inc. proposes to develop a new technology, picoelectrospray ionization mass spectrometry (picoESI-MS), based on our breakthrough multinozzle emitter array, for top-down proteomics of small-volume samples. The technology will be built on our microfabricated monolithic multinozzle emitters (M3 emitters) and multinozzle emitter array (MEA) chips for LC-nanoESI-MS, which collectively offer a straightforward yet novel solution to the longstanding problem of the efficient coupling between silicon microfluidic chips and ESI-MS, and pave the way for the large-scale integration on the proposed microfluidic chips for LC-picoESI-MS. Our picoESI-MS platform will directly address the aforementioned bottlenecks and thus enable high-sensitivity, high-throughput, and multiplex top-down proteomics of small volumes of biological samples.
Public Health Relevance Statement: Public Health Relevance: Cutting-edge technologies enable breakthroughs in biomedical research. Developments of innovative and integrated mass spectrometry-based microfluidic chips will accelerate the discovery and validation of protein biomarkers, thereby providing new strategies for early diagnosis and targeted therapy of diseases.
Project Terms: Address; base; Binding (Molecular Function); Biological; Biological Markers; Biomedical Research; Breast Cancer Cell; Caliber; Charge; Chemistry; Collaborations; Coupling; Detection; Development; Diagnostic; Digestion; Disease; Early Diagnosis; Electrodes; Electrospray Ionization; Human; immunoaffinity chromatography; Individual; Industry; innovation; ionization; Ions; Length; Liquid Chromatography; liquid chromatography mass spectrometry; mass spectrometer; Mass Spectrum Analysis; Measures; Microfluidics; Modification; Molecular Weight; Monitor; new technology; next generation; novel; operation; Pathology; Patients; Peptide Hydrolases; Peptides; Performance; Phase; Physiology; Plasma; programs; Protein Chemistry; Protein Isoforms; protein protein interaction; Proteins; Proteome; Proteomics; public health relevance; Relative (related person); Reproducibility; response; RNA Splicing; Running; Sampling; Sensitivity and Specificity; Shotguns; Silicon; small molecule; Solutions; Solvents; Spectrometry, Mass, Electrospray Ionization; success; Technology; Therapeutic; tool; Trypsin; Validation
